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A molecular switch for stress-induced activation of retrograde mitochondrial transport | 10.1101/2024.09.13.612963 | Gladkova, C.; Paez-Segala, M. G.; Grant, W. P.; Myers, S. A.; Wang, Y.; Vale, R. D. | The cellular distribution of mitochondria in response to stress and local energy needs is governed by the relative activities of kinesin and dynein. The mechanism for switching between these two opposite polarity microtubule motors remains unknown. Here, we coupled a novel cellular synthetic cargo transport assay with AlphaFold2-guided mutagenesis to identify a regulatory helix in the mitochondrial adaptor protein (TRAK) that mediates switching between kinesin- and dynein-driven transport. Differences in the helix sequence explain why two near-identical TRAK isoforms transport mitochondria in predominantly opposite directions. Phosphorylation of the regulatory helix by stress-activated kinases causes the activation of dynein and dissociation of kinesin. Our results reveal a molecular mechanism for coordinating the directional transport of mitochondria in response to intracellular signals. | 2024-09-14 | |
CD11B+CD36+ cells are bone anabolic macrophages that limit age-associated bone loss | 10.1101/2024.09.13.612932 | Koroth, J.; Karkache, I. Y.; Vu, E. K.; Mansky, K. C.; Bradley, E. W. | Disruptions in the bone remodeling cycle that occur with increasing age lead to degeneration of the skeleton and increased risk of fragility fractures. Our understanding of how the bone remodeling process within cortical bone is controlled and altered with age in males and females is limited. Here, we generated bone marrow chimeric mice to understand the impacts of age and sex on the bone remodeling process. We demonstrate that transplantation of aged male or female bone marrow into young lethally irradiated male hosts unexpectedly enhances cortical bone mass without an impacting cancellous bone. Our single cell RNA-sequencing data show that mice reconstituted with aged bone marrow exhibited subsets of cells marked by CD11B/CD36 expression that demonstrate enhanced production of anabolic cytokines as compared to young counterparts, and that these myeloid subsets exist under conditions of normal physiology in aged mice. Importantly, CD11B+CD36+ cells do not differentiate into osteoclasts in vitro, and CD36 does not mark TRAP+ cells in vivo. Instead, CD36+ cells localize to resorption sites, including within cortical bone defects, suggesting their involvement in cortical bone remodeling and healing. CD11B+CD36+ cells also express elevated levels of bone anabolic WNT ligands, especially Wnt6. In functional assays, we demonstrate that soluble factors produced by CD11B+CD36+ cells enhance osteoblast progenitor commitment, mineralization, and activation of WNT signaling in vitro. Moreover, CD11B/CD36 exquisitely mark a subset of anabolic myeloid cells within human bone marrow. In conclusion, our studies identified a novel population of aged macrophages that limit cortical bone loss. | 2024-09-14 | |
CD34+ Stromal Cell/Telocytes Demonstrate a Dynamic Pattern of Distribution During Healing of Post-Infarcted Myocardium in Middle-Aged Sprague-Dawley Rats | 10.1101/2024.09.13.612962 | Schneider, D. T.; Dedkov, E. I. | Introduction: Myocardial CD34+ stromal cells/telocytes (SC/TCs) have been recently recognized as a novel resident cell which may play an important role in the repair process following acute myocardial infarction (MI). This study aims to determine the spatiotemporal dynamics of CD34+ SC/TCs within the left ventricular (LV) wall during the late inflammatory and proliferative phases of post-MI scar formation. Methods: A large transmural MI was induced in middle-aged, Sprague-Dawley rats by permanent ligation of the left anterior descending coronary artery. To recognize proliferating cells, rats were infused with 5-bromo-2'-deoxyuridine (BrdU) in a dose of 12.5 mg/kg/day for 72 hours via intraperitoneal osmotic minipumps on day 0, 4, or 11 after surgery. The rats were euthanized on day 3, 7 and 14 after MI, and their hearts were processed for histology and immunostaining. Results: Three days after MI, CD34+ SC/TCs were absent within the necrotic myocardial tissue but were visible around the surviving cardiac myocytes (CMs) bordering the infarcted region, including those remaining in subepicardial and subendocardial regions, and in the adventitia of residual coronary vessels. Seven days after MI, many of the CD34+ SC/TCs located at the periphery of the developing scar appeared enlarged and contained the BrdU labeling, indicating the cell proliferation. At the same time, elongated CD34+ SC/TCs, which lacked BrdU labeling, were noticed closer to the necrotic zone residing in the interstitial areas between the intact basement membranes left from resorbed CMs, suggesting their migratory activity. Fourteen days after MI, CD34+ SC/TCs were distributed throughout the entire post-infarcted region except for the areas occupied by necrotic tissue, myofibroblast-rich granulation tissue, and the fibroelastic thickenings of the endocardium affected by an MI. Furthermore, accumulated clusters of flattened CD34+ SC/TCs cells were apparent in the areas where the edges of surviving CMs extend into the fibrotic portion of the scar. Conclusion: These findings, for the first time, demonstrate that a population of myocardial CD34+ SC/TCs follow a dynamic pattern of spatiotemporal distribution within the healing myocardium suggesting their direct involvement in post-MI repair process and scar formation. | 2024-09-14 | |
Profiling the physiological impact of aberrant folded-state protein filamentation in cells | 10.1101/2024.09.13.612878 | Levin, T.; Garcia-Seisdedos, H.; Lobov, A.; Wojtynek, M.; Alexandrov, A.; Jona, G.; Levi, D.; Medalia, O.; Levy, E. D. | The formation of large polymeric structures such as cytoskeletal and enzyme filaments is crucial for normal cellular function. However, such filaments can also form due to mutations that create self-interactions at the surface of symmetric proteins. Often, the proteins forming these structures maintain a folded state and thereby differ from aggregates and amyloids that involve misfolding. We refer to this type of assemblies as agglomerates to mark this difference. While cells have quality control mechanisms to identify, buffer, and eliminate misfolded proteins, it is unclear whether similar mechanisms exist for agglomerates, or whether agglomerates are toxic to cells. Here, we profiled the physiological impact of mutation-induced folded-state protein filamentation in yeast cells. First, we devised a simple strategy to distinguish fluorescently labeled proteins forming agglomerates versus aggregates. We then profiled exogenous protein agglomerates in terms of their recognition by known quality control mechanisms, their effects on specific cellular processes and overall fitness on S. cerevisiae cultures. We found that agglomerates do not colocalize with the proteostasis machinery and do not result in measurable fitness defects. Proteomics profiling of cells expressing the wild type protein, agglomerating or misfolded variants revealed a consistent picture, with only minor, agglomerate-size-dependent changes observed and linked to the cell-wall and plasma-membrane proteins. Overall, our findings indicate that agglomerates form mostly benign structures in cells when compared to aggregates, and thereby offer a promising route for synthetic biology applications. | 2024-09-14 | |
Loss of Neurofibromin Induces Inflammatory Macrophage Phenotypic Switch and Retinal Neovascularization via GLUT1 Activation | 10.1101/2024.09.13.612509 | ZAIDI, Y.; Tritz, R.; Zaidi, N.; Nabi, F.; Zaidi, S. A. H.; Morsy, A.; Harris, V.; Racine, R.; Hudson, F. Z.; Bordan, Z.; Kennard, S.; Batori, R.; Huo, Y.; Csanyi, G.; Belin de Chantemele, E. J.; Lei, K.; Boulis, N. M.; Fulton, D. J.; Khan, R. H.; Caldwell, R. B.; Stansfield, B. K. | Persons with neurofibromatosis type 1 (NF1), a tumor predisposition syndrome, are largely protected from diabetes and exhibit evidence of enhanced glucose metabolism, which is replicated in mice harboring Nf1 mutations. A hallmark of NF1-associated neurofibromas and sarcomas is the high density of inflammatory macrophages and targeting macrophages appears efficacious in models of NF1. Inflammatory macrophages rely on glycolysis to rapidly generate ATP; thus, identifying whether neurofibromin, the protein encoded by the NF1 gene, controls glucose uptake and/or glycolysis in macrophages is therapeutically compelling. Using neurofibromin-deficient macrophages and macrophage-specific Nf1 knockout mice, we demonstrate that neurofibromin complexes with glucose transporter 1 (GLUT1) to restrain its activity and that loss of neurofibromin permits Akt2 to facilitate GLUT1 translocation to the membrane in macrophages. In turn, glucose internalization and glycolysis are highly up regulated and provoke putative reparative (M2) macrophages to undergo inflammatory phenotypic switch. Inflammatory M1 macrophages and inflammatory-like M2 macrophages invest the perivascular stroma of tumors and induce pathologic angiogenesis in mice harboring macrophage-specific Nf1 deletion. These studies identify a clear mechanism for the enhanced glycolysis and low risk for diabetes observed in persons with NF1 and provide a novel therapeutic target for manifestations of NF1. | 2024-09-14 | |
Altered polyadenylation site usage in SERPINA1 3'UTR in response to cellular stress affects A1AT protein expression | 10.1101/2024.09.13.612749 | Jiamutai, F.; Hatfield, A.; Herbert, A.; Majumdar, D.; Shankar, V.; Lackey, L. | Alternative polyadenylation results in different 3' isoforms of messenger RNA (mRNA) transcripts. Alternative polyadenylation in the 3' untranslated region (3'UTR) can alter RNA localization, stability and translational efficiency. The SERPINA1 mRNA has two distinct 3' UTR isoforms, both of which express the protease inhibitor -1-antitrypsin (A1AT). A1AT is an acute phase protein that is expressed and secreted from liver hepatocytes and upregulated during inflammation. Low levels of A1AT in the lung contributes to chronic obstructive pulmonary disease, while misfolding of A1AT in the liver contributes to liver cirrhosis. We analyzed the dynamics of alternative polyadenylation during cellular stress by treating the liver cell line HepG2 with the cytokine interleukin 6 (IL-6), ethanol or peroxide. SERPINA1 is transcriptionally upregulated after IL-6 treatment and has altered polyadenylation, resulting in an increase in long 3'UTR isoforms. We find that the long 3'UTR represses endogenous A1AT protein expression even with high levels of SERPINA1 mRNA. SERPINA1 expression and 3' end processing were not affected by ethanol or peroxide. IL-6-induced changes in transcriptome-wide transcriptional regulation suggest changes to the endoplasmic reticulum and in secretory protein processing. Our data suggest that inflammation influences polyA site choice for SERPINA1 transcripts, resulting in reduced A1AT protein expression. | 2024-09-14 | |
Acute sildenafil administration reduces susceptibility to induced atrial fibrillation in sheep | 10.1101/2024.09.09.612118 | denham, N.; Madders, G. W.; Hutchings, D.; Smith, C. E.; Whitley, A.; Obeidat, M.; Trafford, A.; Pearman, C.; Dibb, K. | Background: Sildenafil is a PDE5 inhibitor with a very good safety profile and animal models suggest it may be beneficial in the treatment of heart failure and ventricular fibrillation. Sildenafil has also been associated with a reduced incidence of atrial fibrillation (AF) in a retrospective observational study. We have therefore sought to determine whether sildenafil has a direct effect on atrial electrophysiology and resultant AF burden. Methods: Invasive electrophysiological studies were performed in 12 anaesthetised healthy adult female Welsh mountain sheep. Pacing protocols were performed in the right atrium before and after administration of an acute 10 mg intravenous bolus of sildenafil and the burden of AF assessed. Results: Sildenafil profoundly reduced the vulnerability to AF, decreasing AF duration (112.2 +/- 73.5 s vs. 3.3 +/- 1.4 s), the number of burst pacing inductions causing AF (90 % vs 70 %) and the complexity of AF. The antiarrhythmic effects of sildenafil were determined to be resultant of prolongation of both the atrial effective refractory period (146.9 +/- 7.2 ms vs 166.2 +/- 32.5 ms) and the atrial excitation wavelength (12.9 +/- 0.07 cm vs 15.0 +/- 0.07 cm) and resulted in a shallower restitution curve, reflected in a decreased magnitude of monophasic action potential alternans (0.09 +/- 0.001 mV vs 0.05 +/- 0.10 mV). Conclusions: In the subjectively healthy atria of a highly translational model a strong antiarrhythmic effect upon acute sildenafil application was observed suggestive of a potential clinical benefit in AF. | 2024-09-14 | |
Task-relevant representational formats in multi-layered memory traces | 10.1101/2024.09.10.612205 | Heinen, R.; Rau, E.; Herweg, N.; Axmacher, N. | During encoding, stimuli are embedded into memory traces that allow for their later retrieval. This process is selective, however, because not every aspect of our experiences can be remembered. In addition, post-encoding stages including consolidation are widely assumed to induce transformation processes of the memory trace. It is unclear, however, how selective the memory trace is, whether irrelevant information is completely removed during encoding and/or consolidation, and how this affects retrieval of either general (gist-like) or specific (perceptual) information. Here we show that memory traces consist of multi-layered representational spaces whose formats are flexibly strengthened or weakened during encoding and consolidation depending on task instructions, distinctly shaping their affordances for general or specific retrieval. In a series of behavioral experiments, participants first compared pairs of natural images on either two conceptual or two perceptual dimensions, leading them to incorporate the images into representational spaces defined by Euclidean distances. We found that distances in task-relevant but not irrelevant spaces affected memory strengths. Conceptual encoding benefitted general without impairing specific retrieval, suggesting that perceptual information remained in the memory trace even if it was task-irrelevant. By contrast, targeted memory reactivation (TMR) of conceptual encoding improved memory strength but deteriorated perceptual discrimination during retrieval, indicating that it weakened the accessibility of perceptual formats. Our results demonstrate the flexibility of representational formats that are incorporated into memory traces, and more generally show how the organization of information in representational spaces shapes human behavior. | 2024-09-14 | |
Ethograms reveal a fear conditioned visual cue to organize diverse behaviors in rats | 10.1101/2024.09.10.612214 | Williams, D. C.; Chu, A.; Gordon, N. T.; DuBois, A. M.; Qian, S.; Valvo, G.; Shen, S.; Boyce, J. B.; Fitzpatrick, A. C.; Moaddab, M.; Russell, E. L.; Counsman, L. H.; McDannald, M. A. | Recognizing and responding to threat cues is essential to survival. In rats, freezing is the most common behavior measured. Previously we demonstrated a threat cue can organize diverse behaviors (Chu et al., 2024). However, the experimental design of Chu et al. (2024) was complex and the findings descriptive. Here, we gave female and male Long Evans rats simple paired or unpaired presentations of a light and foot shock (8 total) in a conditioned suppression setting, using a range of shock intensities (0.15, 0.25, 0.35 or 0.5 mA). We found that conditioned suppression was only observed at higher foot shock intensities (0.35 mA and 0.5 mA). We constructed comprehensive, temporal ethograms by scoring 22,272 frames of behavior for 12 mutually exclusive behavior categories in 200 ms intervals around cue presentation. A 0.5 mA and 0.35 mA shock-paired visual cue suppressed reward seeking, rearing and scaling, as well as light-directed rearing and light-directed scaling. The shock-paired visual paired cue further elicited locomotion and freezing. Linear discriminant analyses showed that ethogram data could accurately classify rats into paired and unpaired groups. Considering the complete ethogram data produced superior classification than considering subsets of behaviors. The results demonstrate diverse threat-elicited behaviors - in a simple Pavlovian fear conditioning design - containing sufficient information to distinguish the fear learning status of individual rats. | 2024-09-14 | |
Human microRNA-153-3p targets specific neuronal genes and is associated with the risk of Alzheimer's disease. | 10.1101/2024.09.07.611728 | Wang, R.; Maloney, B.; Nho, K.; Beck, J.; Counts, S. E.; Lahiri, D. K. | Alzheimers disease (AD) is a progressive degenerative disease characterized by a significant loss of neurons and synapses in cognitive brain regions and is the leading cause of dementia worldwide. AD pathology comprises extracellular amyloid plaques and intracellular neurofibrillary tangles. However, the triggers of this pathology are still poorly understood. Repressor element 1-silencing transcription/neuron-restrictive silencer factor (REST/NRSF), a transcription repressor of neuronal genes, is dysregulated during AD pathogenesis. How REST is dysregulated is still poorly understood, especially at the post-transcriptional level. MicroRNAs (miRNAs), a group of short non-coding RNAs, typically regulate protein expression by interacting with target mRNA transcript 3-untranslated region (UTR) and play essential roles in AD pathogenesis. Herein, we demonstrate that miR-153-3p reduces REST 3-UTR activities, mRNA, and protein levels in human cell lines, along with downregulating amyloid {beta} precursor protein (APP) and -synuclein (SNCA). We determine by mutational analyses that miR-153-3p interacts with specific targets via the seed sequence present within the respective mRNA 3-UTR. We show that miR-153-3p treatment alters the expression of these specific proteins in human neuronally differentiated cells and human induced pluripotent stem cells and that miR-153-3p is itself dysregulated in AD. We further find that single nucleotide polymorphisms (SNPs) within 5kb of the MIR153-1 and MIR153-2 genes are associated with AD-related endophenotypes. Elevation of miR-153-3p is associated with reduced AD probability, while elevated REST may associate with a greater AD probability. Our work suggests that a supplement of miR-153-3p would reduce levels of toxic protein aggregates by reducing APP, SNCA, and REST expression, all pointing towards a therapeutic and biomarker potential of miR-153-3p in AD and related dementias. | 2024-09-14 | |
A double hit affecting the IKZF1-IKZF2 tandem in immune cells of schizophrenic patients regulate specific symptoms | 10.1101/2024.09.09.612162 | Ballasch, I.; Lopez-Molina, L.; Galan-Ganga, M.; Sancho-Balsells, A.; Rodriguez-Navarro, I.; Borras-Pernas, S.; Rabadan, M.-A.; Chen, W.; Pasto-Pellicer, C.; Flotta, F.; Fernandez-Irigoyen, J.; Santamaria, E.; Aguilar, R.; Dobano, C.; Egri, N.; Hernandez, C.; Alfonso, M.; Juan, M.; Alberch, J.; del Toro, D.; Arranz, B.; Canals, J.-M.; Giralt, A. | Schizophrenia is a complex multifactorial disorder and increasing evidence suggests the involvement of immune dysregulations in its pathogenesis. We observed that IKZF1 and IKZF2, classic immune-related transcription factors (TFs), were both downregulated in patients peripheral blood mononuclear cells (PBMCs) but not in their brain. We generated a new mutant mouse model with a reduction in Ikzf1 and Ikzf2 to study the impact of those changes. Such mice developed deficits in the three dimensions (positive-negative-cognitive) of schizophrenic-like phenotypes associated with alterations in structural synaptic plasticity. We then studied the secretomes of cultured PBMCs obtained from human patients and identified potentially secreted molecules, which depended on IKZF1 and IKZF2 levels, and that in turn have an impact on neural synchrony, structural synaptic plasticity and schizophrenic-like symptoms in in vivo and in vitro models. Our results point out that IKZF1-IKZF2-dependent immune signals negatively impact on essential neural circuits involved in schizophrenia. | 2024-09-14 | |
Epigenetic adaptation drives monocyte differentiation into microglia-like cells upon engraftment into the retina | 10.1101/2024.09.09.612126 | Liu, J.; Lei, F.; Yan, B.; Cui, N.; Sharma, J.; Correa, V.; Roach, L.; Kikolaou, S.; Pitts, K.; Chodosh, J.; Maidana, D. E.; Vavvas, D. G.; Zhang, H.; Weitz, D.; Mostoslavsky, R.; Paschalis, E. I. | The identification of specific markers for microglia has been a long-standing challenge. Recently, markers such as P2ry12, TMEM119, and Fcrls have been proposed as microglia-specific and widely used to explore microglial functions within various central nervous system (CNS) contexts. The specificity of these markers was based on the assumption that circulating monocytes retain their distinct signatures even after infiltrating the CNS. However, recent findings reveal that infiltrating monocytes can adopt microglia-like characteristics while maintaining a pro-inflammatory profile upon permanent engraftment in the CNS.In this study, we utilize bone marrow chimeras, single-cell RNA sequencing, ATAC-seq, flow cytometry, and immunohistochemistry to demonstrate that engrafted monocytes acquire expression of established microglia markers P2ry12, TMEM119, Fcrls and the pan-myeloid marker Iba1, which has been commonly mischaracterized as microglia-specific. These changes are accompanied by alterations in chromatin accessibility and shifts in chromatin binding motifs that are indicative of microglial identity. Moreover, we show that engrafted monocytes dynamically regulate the expression of CX3CR1, CCR2, Ly6C, and transcription factors PU.1, CTCF, RUNX, AP-1, CEBP, and IRF2, all of which are crucial for shaping microglial identity. This study is the first to illustrate that engrafted monocytes in the retina undergo both epigenetic and transcriptional changes, enabling them to express microglia-like signatures. These findings highlight the need for future research to account for these changes when assessing the roles of monocytes and microglia in CNS pathology. | 2024-09-14 | |
Synthetic G-quadruplex components for predictable, precise two-level control of mammalian recombinant protein expression | 10.1101/2024.09.10.612233 | Pohle, M.; Curry, E.; Gibson, S.; Brown, A. | Control of mammalian recombinant protein expression underpins the in vitro manufacture and in vivo performance of all biopharmaceutical products. However, routine optimization of protein expression levels in these applications is hampered by a paucity of genetic elements that function predictably across varying molecular formats and host cell contexts. Herein, we describe synthetic genetic components that are specifically built to simplify bioindustrial expression cassette design processes. Synthetic G-quadruplex elements with varying sequence feature compositions were systematically designed to exhibit a wide-range of regulatory activities, and inserted into identified optimal positions within a standardized, bioindustry compatible core promoter-5' UTR control unit. The resulting library tuned protein production rates over two orders of magnitude, where DNA and RNA G-quadruplexes could be deployed individually, or in combination to achieve synergistic two-level regulatory control. We demonstrate these components can predictably and precisely tailor protein expression levels in i) varying gene therapy and biomanufacturing cell hosts, and ii) both plasmid DNA and synthetic mRNA contexts. As an exemplar use-case, a vector design platform was created to facilitate rapid optimization of polypeptide expression ratios for difficult-to-express multichain products. Permitting simple, predictable titration of recombinant protein expression, this technology should prove useful for gene therapy and biopharmaceutical manufacturing applications. | 2024-09-14 | |
Mechanisms of peptide agonist dissociation and deactivation of adhesion G-protein-coupled receptors | 10.1101/2024.09.07.611823 | Joshi, K.; Miao, Y. | Adhesion G protein coupled receptors (ADGRs) belong to Class B2 of GPCRs and are involved in a wide array of important physiological processes. ADGRs contain a GPCR autoproteolysis-inducing (GAIN) domain that is proximal to the receptor N-terminus and undergoes autoproteolysis during biosynthesis to generate two fragments: the N-terminal fragment (NTF) and C-terminal fragment (CTF). Dissociation of NTF reveals a tethered agonist to activate CTF of ADGRs for G protein signaling. Synthetic peptides that mimic the tethered agonist can also activate the ADGRs. However, mechanisms of peptide agonist dissociation and deactivation of ADGRs remain poorly understood. In this study, we have performed all-atom enhanced sampling simulations using a novel Protein Protein Interaction-Gaussian accelerated Molecular Dynamics (PPI GaMD) method on the ADGRG2 IP15 and ADGRG1 P7 complexes. The PPI GaMD simulations captured dissociation of the IP15 and P7 peptide agonists from their target receptors. We were able to identify important low-energy conformations of ADGRG2 and ADGRG1 in the active, intermediate, and inactive states, as well as exploring different states of the peptide agonists IP15 and P7 during dissociation. Therefore, our PPI-GaMD simulations have revealed dynamic mechanisms of peptide agonist dissociation and deactivation of ADGRG1 and ADGRG2, which will facilitate rational design of peptide regulators of the two receptors and other ADGRs. Keywords: adhesion G protein-coupled receptors; Protein-Protein Interaction-Gaussian accelerated Molecular Dynamics; enhanced sampling, peptide dissociation; deactivation. | 2024-09-14 | |
Primary cilia promote EMT-induced triple-negative breast tumor heterogeneity and resistance to therapy | 10.1101/2024.09.10.612175 | Tessier, C.; Derrien, J.; Dupuy, A.; Pele, T.; Moquet, M.; Roul, J.; Douillard, E.; El Harrif, C.; Pinson, X.; Le Gallo, M.; Godey, F.; Tas, P.; Viel, R.; Prigent, C.; Letouze, E.; Suzanne, P.; Dallemagne, P.; Campone, M.; Weinberg, R.; Lees, J.; Juin, P.; Guen, V. | Tumor heterogeneity and plasticity, driven by Epithelial-Mesenchymal Transition (EMT), enable cancer therapeutic resistance. We previously showed that EMT promotes primary cilia formation, which enables stemness and tumorigenesis in triple-negative breast cancer (TNBC). Here, we establish a role for primary cilia in human TNBC chemotherapeutic resistance. We developed patient-derived organoids, and showed that these recapitulated the cellular heterogeneity of TNBC biopsies. Notably, one of the identified cell states bore a quasi-mesenchymal phenotype, primary cilia, and stemness signatures. We treated our TNBC organoids with chemotherapeutics and observed partial killing. The surviving cells with organoid-reconstituting capacity showed selective enrichment for the quasi-mesenchymal ciliated cell subpopulation. Genomic analyses argue that this enrichment reflects a combination of pre-existing cells and ones that arose through drug-induced cellular plasticity. We developed a family of small-molecule inhibitors of ciliogenesis and show that these, or genetic ablation of primary cilia, suppress chemoresistance. We conclude that primary cilia help TNBC to evade chemotherapy. | 2024-09-14 | |
NEK9 ablation rewires docetaxel resistance through induction of ERK-mediated cancer cell pyroptosis | 10.1101/2024.09.10.612209 | Ansari, S. A.; Mohanty, S.; Mohapatra, P.; Rath, R.; Muduli, D.; Majumdar, S. K. D.; Swain, R. K.; Dash, R. | Docetaxel alone or in combination with other drugs is the most common chemotherapy regimen for several neoplasms including advanced OSCC. Unfortunately, chemoresistance leads to relapse and continued tumor growth. It is therefore important to explore the causative factors for docetaxel resistance. In this study, we performed a CRISPR-based kinome screening that identified Never In Mitosis Gene-A Related Kinase-9 (NEK9) as a major player of docetaxel resistance in OSCC, prostate, and pancreatic cancer lines. NEK9 expression was upregulated in tumor samples of chemotherapy non-responders compared to responder OSCC patients. Our validation data suggests selectively knocking out NEK9 sensitizes cancer cells to docetaxel. Mechanistically, we found that ablation of NEK9 induces DNA damage, activating ERK(p-T202/Y204) that leads to Gasdermin-E mediated Cancer Cell pyroptosis. The in-vitro kinase activity assay identified fostamatinib as a potent inhibitor of NEK9. The xenograft data suggest that fostamatinib restores docetaxel sensitivity and facilitates a significant reduction of tumor burden. Overall, our data suggests a novel combination of fostamatinib and docetaxel needs further clinical investigation in advanced OSCC. | 2024-09-14 | |
RAD50 is a potential biomarker for breast cancer diagnosis and prognosis | 10.1101/2024.09.07.611821 | Chhatwal, K. S.; Liu, H. | BACKGROUND: RAD50 is one of the most critical genes in DNA double-strand break processing, which can lead to a single strand of DNA overhang and is potentially involved in forcing incomplete DNA repair. This research study aims to investigate the role of RAD50 in breast cancer diagnosis and prognosis. METHODS: Breast cancer mRNA expression data was collected from TCGA and the difference between cancer and non-cancer in gene expression of RAD50 was analyzed. The survival association of RAD50 was also analyzed. RESULTS: RAD50 expression is significantly lower in cancer than in normal tissue. High expression of RAD50 is associated with worse survival. Conclusion: RAD50 is a potential biomarker for breast cancer diagnosis and prognosis | 2024-09-14 | |
Unlocking expanded flagellin perception through rational receptor engineering | 10.1101/2024.09.09.612155 | Li, T.; Bolanos, E. J.; Stevens, D.; Sha, H.; Prigozhin, D. M.; Coaker, G. | The surface-localized receptor kinase FLS2 detects the flg22 epitope from bacterial flagella. FLS2 is conserved across land plants, but bacterial pathogens exhibit polymorphic flg22 epitopes. Most FLS2 homologs possess narrow perception ranges, but four with expanded perception ranges have been identified. Using diversity analyses, AlphaFold modeling, and amino acid properties, key residues enabling expanded recognition were mapped to FLS2's concave surface, interacting with the co-receptor and polymorphic flg22 residues. Synthetic biology enabled engineering of expanded recognition from QvFLS2 (Quercus variabilis) and FLS2XL (Vitis riparia) into homologs with canonical perception. Evolutionary analyses across three plant orders showed residues under positive selection aligning with those binding the co-receptor and flg22's C-terminus, suggesting more alleles with expanded perception exist. Our experimental data enabled the identification of specific receptor amino acid properties and AlphaFold3 metrics that facilitate predicting FLS2-flg22 recognition. This study provides a framework for rational receptor engineering to enhance pathogen restriction. | 2024-09-14 | |
Sex-dependent effects of a high-fat diet on the hypothalamic response in mouse | 10.1101/2024.09.10.612192 | Dreux, V.; Lefebvre, C.; Breemeersch, C.-E.; Salaun, C.; Bole-Feysot, C.; Guerin, C.; Dechelotte, P.; Goichon, A.; Coeffier, M.; Langlois, L. D. | Sex differences in rodent models of diet-induced obesity are still poorly documented, particularly regarding how central mechanisms vary between sexes in response to an obesogenic diet. Here, we wanted to determine whether obesity phenotype and hypothalamic response differed between male and female C57Bl/6J mice when exposed to a high-fat diet (HFD). Mice were exposed either a free 60% HFD or standard diet first for both a long- (14 weeks) and shorter-periods of time (3, 7, 14 and 28 days). Analysis of the expression profile of key neuronal, glial and inflammatory hypothalamic markers was performed using RT-qPCR. In addition, astrocytic and microglial morphology was examined in the arcuate nucleus. Monitoring of body weight and composition revealed that body weight and fat mass gain appeared earlier and was more pronounced in male mice. After 14 weeks of HFD exposure, normalized increase of body weight reached similar levels between male and female mice. Overall, both sexes under HFD displayed a decrease of orexigenic neuropeptides expression and an increase in POMC gene expression was observed only in female mice. In addition, changes in the expression of hypothalamic inflammatory markers were relatively modest. We also reported that the glial cell markers expression and morphology were affected by HFD in a sex- and time dependent manner, suggesting a more pronounced glial cell activation in female mice. Taken together, these data show that male and female mice responded differently to HFD exposure, both on short- and long-term and suggest that a strong inflammatory hypothalamic profile is not systematically present in DIO models. Nevertheless, in addition to these present data, the underlying mechanisms should be deciphered in further investigations. | 2024-09-14 | |
Wakefulness induced by TAAR1 partial agonism is mediated through dopaminergic neurotransmission | 10.1101/2024.09.09.612122 | Park, S.; Heu, J.; Hoener, M. C.; Kilduff, T. S. | Trace amine-associated receptor 1 (TAAR1) is a negative regulator of dopamine (DA) release. The partial TAAR1 agonist RO5263397 promotes wakefulness and suppresses NREM and REM sleep in rodents and non-human primates. We tested the hypothesis that the TAAR1-mediated effects on sleep/wake were due, in part, to DA release. Male C57BL6/J mice (n=8) were intraper-itoneally administered the D1R antagonist SCH23390, the D2R antagonist eticlopride, a combi-nation of D1R+D2R antagonists or saline at ZT5.5, followed 30 min later by RO5263397 or vehi-cle per os. EEG, EMG, subcutaneous temperature, and activity were recorded across the 8 treat-ments and sleep architecture was analyzed for 6 hours post-dosing. As described previously, RO5263397 increased wakefulness and delayed NREM and REM sleep onset. D1, D2, and D1+D2 pretreatment reduced RO5263397-induced wakefulness for 1-2 hours after dosing but only the D1 antagonist significantly reduced the TAAR1-mediated increase in NREM latency. Neither the D1 nor the D2 antagonist affected TAAR1-mediated suppression of REM sleep. These results suggest that, whereas TAAR1 effects on wakefulness are mediated in part through the D2R, D1R activation plays a role in reversing the TAAR1-mediated increase in NREM sleep latency. By contrast, TAAR1-mediated suppression of REM sleep appears not to involve D1R or D2R mechanisms. | 2024-09-14 | |
Ultrastructural analysis of the brain endothelium by electron tomography | 10.1101/2024.09.09.612128 | Kotchetkov, P.; Lacoste, B. | Transmission electron microscopy (TEM) is a powerful imaging technique, yielding ultrastructural investigation of organic and non-organic samples. Despite its ability to reach nanoscale resolutions, conventional TEM presents a major disadvantage by only acquiring two-dimensional snapshots, thus hindering our volumetric understanding of samples. Electron tomography (ET) overcomes this limitation by offering detailed views of a thin specimen in 3 dimensions (3D). This technique is widely used in biology and has expanded our understanding of mitochondrial structure or synaptic organization. Proper brain functioning is highly reliable on a constant nutritional support through its microvasculature lined by endothelial cells. These unique cells form a selective and protective barrier, known as the blood-brain barrier (BBB), which limits the entrance of blood-borne molecules into the brain. In pathological conditions, the BBB is disrupted, resulting in neuronal damage. Understanding the fine changes underlying BBB disruption requires advanced imaging tools such as ET, to detect the finest changes in endothelial ultrastructure. This manuscript briefly explains how TEM and ET function, and then provides a detailed, didactic method for sample preparation, tomogram generation and 3D segmentation of brain endothelial cells using ET. | 2024-09-14 | |
A Statistically-Robust Model Of The Axomyelin Unit Under Normal Physiologic Conditions With Application To Disease States | 10.1101/2024.09.10.612208 | Gow, A.; Dupree, J. L.; Feinstein, D.; Boullerne, A. | Despite tremendous progress in characterizing the myriad cellular structures in the nervous system, a full appreciation of the interdependent and intricate interactions between these structures is as yet unfulfilled. Indeed, few more so than the interaction between the myelin internode and its ensheathed axon. More than a half-century after the ultrastructural characterization of this axomyelin unit, we lack a reliable understanding of the physiological properties, the significance and consequence of pathobiological processes, and the means to gauge success or failure of interventions designed to mitigate disease. Herein, we highlight shortcomings in the most common statistical procedures used to characterize the axomyelin unit, with particular emphasis on the underlying principles of simple linear regression. These shortcomings lead to insensitive detection and/or ambiguous interpretation of normal physiology, disease mechanisms and remedial methodologies. To address these problems, we syndicate insights from early seminal myelin studies and use a statistical model of the axomyelin unit that is established in the accompanying article. Herein, we develop and demonstrate a statistically-robust analysis pipeline with which to examine and interpret axomyelin physiology and pathobiology in two disease states, experimental autoimmune encephalomyelitis and the rumpshaker mouse model of leukodystrophy. On a cautionary note, our pipeline is a relatively simple and streamlined approach that is not necessarily a panacea for all g ratio analyses. Rather, it approximates a minimum effort needed to elucidate departures from normal physiology and to determine if more comprehensive studies may lead to deeper insights. | 2024-09-14 | |
Modulation of cortico-muscular coupling associated with split-belt locomotor adaptation | 10.1101/2024.09.09.612154 | Oshima, A.; Yokoyama, H.; Kaneko, N.; Takahashi, R.; Takiyama, K.; Nakazawa, K. | Humans can adjust their walking patterns according to the demands of their internal and external environments, referred to as locomotor adaptation. Although significant functional coupling (i.e. cortico-muscular coherence [CMC]) has been shown between cortical and lower-limb muscle activity during steady-state walking, little is known about CMC in locomotor adaptation. Therefore, we investigated the adaptation-dependent modulation of the CMC between the sensorimotor region and the tibialis anterior muscle using a split-belt locomotor adaptation paradigm that can impose an asymmetric perturbation. We hypothesized that the CMC would temporarily decrease after exposure to the asymmetric perturbation and removal of the perturbation because of a mismatch between the predicted and actual sensory feedback. We also hypothesized that the CMC would increase as adaptation and de-adaptation to perturbation progressed because the motor system could become able to predict sensory feedback. Our findings revealed that the CMC temporarily decreased after exposure to and removal of the perturbation. Moreover, the CMC increased with adaptation and de-adaptation to perturbation. Although these results depend on the leg, frequency bands, and gait phases, they partially support our hypothesis. These findings suggest that flexible updating of cortico-muscular coupling in the motor system is a key mechanism underlying locomotor adaptation in humans. The results from our study on healthy young individuals contribute to the understanding of neuromuscular control of gait and provide valuable insight for optimising gait rehabilitation. | 2024-09-14 | |
Stab Wound Injury Elicits Transit Amplifying Progenitor-like Phenotype in Parenchymal Astrocytes | 10.1101/2024.09.10.612217 | Ninkovic, J.; Maddhesiya, P.; Koupourtidou, C.; Zambusi, A.; Novoselc, K. T.; Fischer-Sternjak, J.; Simon, T.; Olpe, C.; Jessberger, S.; Goetz, M. | Astrocytes exhibit dual roles in central nervous system (CNS) recovery, offering both beneficial and detrimental effects. Following CNS injury, a subset of astrocytes undergoes proliferation, de-differentiation, and acquires self-renewal and neurosphere-forming capabilities in vitro. This subset of astrocytes represents a promising target for initiating brain repair processes and holds potential for neural recovery. However, studying these rare plastic astrocytes is challenging due to the absence of distinct markers. In our study, we characterized these astrocytic subpopulations using comparative single-cell transcriptome analysis. By leveraging the regenerative properties observed in radial glia of zebrafish, we identified and characterized injury-induced plastic astrocytes in mice. These injury-induced astrocytic subpopulations were predominantly proliferative and demonstrated the capacity for self-renewal and neurosphere formation, ultimately differentiating exclusively into astrocytes. Integration with scRNAseq data of the subependymal zone (SEZ) allowed us to trace the origins of these injury-induced plastic astrocytic subpopulations to parenchymal astrocytes. Our analysis revealed that a subset of these injury-induced astrocytes shares transcriptional similarities with endogenous transient amplifying progenitors (TAPs) within the SEZ, rather than with neural stem cells (NSCs). Notably, these injury-induced TAP-like cells exhibit distinct differentiation trajectories, favoring gliogenic over neurogenic differentiation. In summary, our study identifies a rare subset of injury-induced, proliferative plastic astrocytes with neurosphere-forming capacities. These cells originate from reactive astrocytes and resemble TAPs in their transcriptional profile. This study enhances our understanding of astrocyte plasticity post-injury. | 2024-09-14 | |
Prediction of Protein Half-lives from Amino Acid Sequences by Protein Language Models | 10.1101/2024.09.10.612367 | Sagawa, T.; Kanao, E.; Ogata, K.; Imami, K.; Ishihama, Y. | We developed a protein half-life prediction model, PLTNUM, based on a protein language model using an extensive dataset of protein sequences and protein half-lives from the NIH3T3 mouse embryo fibroblast cell line as a training set. PLTNUM achieved an accuracy of 71% on validation data and showed robust performance with an ROC of 0.73 when applied to a human cell line dataset. By incorporating Shapley Additive Explanations (SHAP) into PLTNUM, we identified key factors contributing to shorter protein half-lives, such as cysteine-containing domains and intrinsically disordered regions. Using SHAP values, PLTNUM can also predict potential degron sequences that shorten protein half-lives. This model provides a platform for elucidating the sequence dependency of protein half-lives, while the uncertainty in predictions underscores the importance of biological context in influencing protein half-lives. | 2024-09-14 | |
A new framework for SubtiWiki, the database for the model organism Bacillus subtilis | 10.1101/2024.09.10.612211 | Elfmann, C.; Dumann, V.; van den Berg, T.; Stulke, J. | Bacillus subtilis is a Gram-positive model bacterium and one of the most-studied and best understood organisms. The complex information resulting from its investigation is compiled in the database SubtiWiki (https://subtiwiki.uni-goettingen.de/v5) in an integrated and intuitive manner. To enhance the utility of SubtiWiki, we have added novel features such as a viewer to interrogate conserved genomic organization, a widget that shows mutant fitness data for all non-essential genes, and a widget showing protein structures, structure predictions and complex structures. Moreover, we have integrated metabolites as new entities. The new framework also includes a documented API, enabling programmatic access to data for computational tasks. Here we present the recent developments of SubtiWiki and the current state of the data for this organism. | 2024-09-14 | |
Identifying, Prioritizing, and Visualizing Functional Promoter SNVs with the Recurrence-agnostic REMIND-Cancer Pipeline and pSNV Hunter | 10.1101/2024.09.10.612212 | Abad, N. A. B.; Glas, I.; Hong, C.; Pageaud, Y.; Hutter, B.; Brors, B.; Korner, C.; Feuerbach, L. | Cancer is a heterogeneous disease that arises due to mutations that drive cancer progression. However, the identification of these functional mutations has typically focused only on protein-coding DNA. Among non-coding mutations, only a few have been clearly associated with cancer. We hypothesize that this gap in discovery is partly due to the limitations of current methods requiring high recurrence of mutations. To support candidate selection for experimental validation of lowly recurrent and singleton promoter mutations, new computational approaches for the integrated analysis of multi-omics data are required. To address this challenge, the REMIND-Cancer Pipeline leverages whole-genome sequencing and RNA-Seq data to extract and prioritize functional promoter mutations, regardless of their recurrence status. Subsequently, pSNV Hunter aggregates and visualizes comprehensive information for each candidate. We demonstrate the functionality of both tools by applying it to the PCAWG dataset. This workflow successfully identified and prioritized known highly-recurrent mutations, as well as, novel singletons and lowly recurrent candidates. Hence, the output of our workflow directly supports hypothesis generation for subsequent experimental validation to overcome limitations of recurrence-based approaches. | 2024-09-14 | |
Targeting the Schwann Cell EP2/cAMP Nanodomain to Block Pain but not Inflammation | 10.1101/2024.09.10.612200 | Nassini, R.; Landini, L.; Marini, M.; Chieca, M.; Souza Monteiro de Araujo, D.; Montini, M.; Pensieri, P.; Abruzzese, V. D.; De Siena, G.; Zhang, J.; De Giorgi, V.; Romitelli, A.; Brancolini, G.; Tonello, R.; Peach, C. J.; Mastricci, A.; Scuffi, I.; Tesi, M.; Jensen, D. D.; Schmidt, B. L.; Bunnett, N. W.; De Logu, F.; Geppetti, P. | Analgesia by non-steroidal anti-inflammatory drugs (NSAIDs) is ascribed to inhibition of prostaglandin (PG) biosynthesis and ensuing inflammation. However, NSAIDs have life-threatening side effects, and inhibition of inflammation delays pain resolution. Decoupling the mechanisms underlying PG-evoked pain vs. protective inflammation would facilitate pain treatment. Herein, we reveal that selective silencing of the PGE2 EP2 receptor in Schwann cells via an adeno-associated viral vector abrogates the indomethacin-sensitive component of pain-like responses in mice elicited by inflammatory stimuli without affecting inflammation. In human Schwann cells and in mice, EP2 activation and optogenetic stimulation of adenylyl cyclase evokes a plasma membrane-compartmentalized cyclic adenosine monophosphate (cAMP) signal that, via A-kinase anchor protein-associated protein kinase A, sustains inflammatory pain-like responses, but does not delay their resolution. Thus, an unforeseen and druggable EP2 receptor in Schwann cells, via specific cAMP nanodomains, encodes PG-mediated persistent inflammatory pain but not protective inflammation. | 2024-09-14 | |
Nanomedicines targeting signaling of protease-activated receptor 2 in organelles provide sustained analgesia | 10.1101/2024.09.10.612022 | Teng, S. L.; Latorre, R.; Bhansali, D.; Lewis, P. K.; Pollard, R. E.; Peach, C. J.; Sokrat, B.; Thanigai, G. S. A.; Chiu, T.; Jensen, D. D.; Jimenez-Vargas, N. N.; Mocherniak, A.; Parreiras-E-Silva, L. T.; Bouvier, M.; Bogyo, M.; Gaspari, M. M.; Vanner, S. J.; Pinkerton, N. M.; Leong, K. W.; Schmidt, B. L.; Bunnett, N. W. | Although many internalized G protein-coupled receptors (GPCRs) continue to signal, the mechanisms and outcomes of GPCR signaling in organelles are uncertain due to the challenges of measuring organelle-specific signals and of selectively antagonizing receptors in intracellular compartments. Herein, genetically-encoded biosensors targeted to subcellular compartments were used to analyze organelle-specific signaling of protease-activated receptor 2 (PAR2); the propensity of nanoparticles (NPs) to accumulate in endosomes was leveraged to selectively antagonize intracellular PAR2 signaling of pain. PAR2 agonists evoked sustained activation of PAR2, Gq and {beta}-arrestin-1 in early, late and recycling endosomes and the cis- and trans. Golgi apparatus, and activated extracellular signal regulated kinase (ERK) in the cytosol and nucleus, measured with organelle-targeted biosensors. Dendrimer and core-shell polymeric NPs accumulated in early and late endosomes of HEK293 cells, colonic epithelial cells and nociceptors, detected by confocal imaging of fluorescent NPs. NPs efficiently encapsulated and slowly released AZ3451, a negative allosteric PAR2 antagonist. NP-encapsulated AZ3451, but not unencapsulated AZ3451, rapidly and completely reversed PAR2, Gq and {beta}-arrestin-1 activation in endosomes and the Golgi apparatus and ERK activation in the cytosol and nucleus. When administered into the mouse colon lumen, dendrimer NPs accumulated in endosomes of colonocytes and polymeric NPs targeted neurons, sites of PAR2 expression. Both NP-AZ3451 formulations, but not unencapsulated AZ3451, caused long-lasting analgesia and normalized aberrant behavior in preclinical models of inflammatory bowel disease. Thus, organelle-specific PAR2 signals in colonocytes and nociceptors mediate pain. Antagonism of PAR2 in organelles, rather than at the plasma membrane, provides effective pain relief. | 2024-09-14 | |
Sequential design of single-cell experiments to identify discrete stochastic models for gene expression. | 10.1101/2024.09.12.612709 | Cook, J.; Ron, E.; Svetlov, D.; Aguiulera, L.; Munsky, B. | Control of gene regulation requires quantitatively accurate predictions of heterogeneous cellular responses. When inferred from single-cell experiments, discrete stochastic models can enable such predictions, but such experiments are highly adjustable, allowing for almost infinitely many potential designs (e.g., at different induction levels, for different measurement times, or considering different observed biological species). Not all experiments are equally informative, experiments are time-consuming or expensive to perform, and research begins with limited prior information with which to construct models. To address these concerns, we developed a sequential experiment design strategy that starts with simple preliminary experiments and then integrates chemical master equations to compute the likelihood of single-cell data, a Bayesian inference procedure to sample posterior parameter distributions, and a finite state projection based Fisher information matrix to estimate the expected information for different designs for subsequent experiments. Using simulated then real single-cell data, we determined practical working principles to reduce the overall number of experiments needed to achieve predictive, quantitative understanding of single-cell responses. | 2024-09-14 | |
Revealing the neural representations underlying other-race face perception | 10.1101/2024.09.09.611889 | Shoura, M.; Liang, Y. Z.; Sama, M.; Nestor, A. | The other-race effect, a disadvantage at recognizing faces of other races than one's own, has received considerable attention, especially regarding its wide scope and underlying mechanisms. Here, we aim to elucidate its neural and representational basis by relating behavioral performance in East Asian and White individuals to neural decoding and image reconstruction relying on electroencephalography data. Our investigation uncovers a reliable neural counterpart of the other-race effect (i.e., a decoding disadvantage for other-race faces) along with its extended dynamics and prominence across individuals. Further, it retrieves, via neural-based image reconstruction, visual representations underlying other-race face perception and their intrinsic biases. Notably, our data-driven approach reveals that other-race faces are perceived not just as more typical but, also, as younger and more expressive. These findings, pointing to multiple visual biases surrounding the other-race effect, speak to the complexity of its neural mechanisms and its social implications. | 2024-09-14 | |
A brain-body feedback loop driving HPA-axis dysfunction in breast cancer | 10.1101/2024.09.13.612923 | Gomez, A.; Wu, Y.; Zhang, C.; Boyd, L.; Wee, T. L.; Gewolb, J.; Amor, C.; Cheadle, L. M.; Borniger, J. C. | Breast cancer patients often exhibit disrupted circadian rhythms in circulating glucocorticoids (GCs), such as cortisol. This disruption correlates with reduced quality of life and higher cancer mortality. The exact cause of this phenomenon; whether due to treatments, stress, age, co-morbidities, lifestyle factors, or the cancer itself remains unclear. Here, we demonstrate that primary breast cancer alone blunts host GC rhythms by disinhibiting neurons in the hypothalamus, and that circadian phase-specific neuromodulation of these neurons can attenuate tumor growth by enhancing anti-tumor immunity. We find that mice with mammary tumors exhibit blunted GC rhythms before tumors are palpable, alongside increased activity in paraventricular hypothalamic neurons expressing corticotropin-releasing hormone (i.e., PVNCRH neurons). Tumor-bearing mice have fewer inhibitory synapses contacting PVNCRH neurons and reduced miniature inhibitory post-synaptic current (mIPSC) frequency, leading to net excitation. Tumor-bearing mice experience impaired negative feedback on GC production, but adrenal and pituitary gland functions are largely unaffected, indicating that alterations in PVNCRH neuronal activity are likely a primary cause of hypothalamic-pituitary-adrenal (HPA) axis dysfunction in breast cancer. Using chemogenetics (hM3Dq) to stimulate PVNCRH neurons at different circadian phases, we show that stimulation just before the light-to-dark transition restores normal GC rhythms and reduces tumor progression. These mice have significantly more effector T cells (CD8+) within the tumor than non-stimulated controls, and the anti-tumor effect of PVNCRH neuronal stimulation is absent in mice lacking CD8+ T cells. Our findings demonstrate that breast cancer distally regulates neurons in the hypothalamus that control output of the HPA axis and provide evidence that therapeutic targeting of these neurons could mitigate tumor progression. | 2024-09-14 | |
TAAR2-9 Knockout Mice Exhibit Reduced Wakefulness and Disrupted REM Sleep | 10.1101/2024.09.09.612114 | Park, S.; Heu, J.; Scheldrup, G.; Tisdale, R.; Sun, Y.; Haire, M.; Ma, S.-C.; Hoener, M. C.; Kilduff, T. S. | Trace amine-associated receptor 1 (TAAR1) has gained attention for its roles in modulating neural systems, sleep/wake control, and as a therapeutic target for neuropsychiatric disorders. Although TAARs 2-9 were initially identified as non-canonical olfactory receptors, recent studies have identified extra-nasal receptor distribution of multiple TAARs. To evaluate whether TAARs 2-9 have a role in arousal state regulation, we investigated sleep/wake control in male TAAR2-9 knockout (KO) mice. After determination of baseline sleep/wake patterns, the homeostatic response to sleep deprivation and response to TAAR1 agonists were compared between KO and C57BL/6J mice. Although the EEG of TAAR2-9 KO mice had lower power in the delta and theta bands and higher power in the gamma range, sleep/wake states were readily identified. KO mice had more NREM sleep during the dark phase and more REM sleep during the light phase. Sleep/wake was fragmented in KO mice with shorter Wake and REM bouts during the dark phase and more REM bouts during the light phase. KO mice exhibited more REM sleep during a sleep latency test but the homeostatic response to sleep loss did not differ between the strains. A high dose of the TAAR1 agonist RO5256390 increased Wake and reduced NREM sleep in KO mice whereas RO5256390 and the partial TAAR1 agonist RO5263397 suppressed REM sleep. The number of tyrosine hydroxylase-immunoreactive neurons in the ventral tegmental area was significantly elevated in KO mice. These dopaminergic and sleep/wake alterations in TAAR2-9 KO mice highlight the need for further elucidation of the functions of TAAR2-9. | 2024-09-14 | |
The cardiac, respiratory and gastric rhythms independently modulate corticospinal excitability | 10.1101/2024.09.10.612221 | Engelen, T.; Schuhmann, T.; Sack, A. T.; Tallon-Baudry, C. | Interoception refers to the sensing of the internal state of the body and encompasses various bodily axes. Yet many interoceptive signals display unique qualities. The heart, lungs, and stomach each have their distinct frequencies, afferent pathways, and respective functions. At the same time each of these organs has been demonstrated to interact with neural activity and behaviour. To what extent then should different organs be treated as separate modalities in interoception? We here aim to answer this question by assessing in human participants whether the phase of these visceral rhythms is coupled to the corticospinal excitability of the motor system, and whether this coupling happens in an organ-specific or organ-general manner. We combined continuous physiological recordings with single pulse Transcranial Magnetic Stimulation (TMS) to probe phase-amplitude coupling between the phase of the cardiac, respiratory, and gastric rhythm and the amplitude of Motor Evoked Potentials (MEP). All three visceral rhythms contributed to MEP amplitude with similar effect sizes at the group level. However, we found no relation between coupling strengths with corticospinal excitability between the three organs. Thus, participants displaying high coupling with one organ did not necessarily display high coupling to the other organs, suggestive of unique interoceptive profiles. There was also no link between self-reported awareness of the organ and the actual coupling, suggesting these are distinct dimensions of interoception. Together these results show that each coupling is mediated by at least partially independent mechanisms. | 2024-09-14 | |
Parechovirus-3 infection disrupts immunometabolism and leads to glutamate excitotoxicity in neural organoids | 10.1101/2024.09.10.611955 | Capendale, P. E.; Ambikan, A. T.; Garcia Rodriguez, I.; Vieira de Sa, R.; Pajkrt, D.; Wolthers, K. C.; Neogi, U.; Sridhar, A. | Parechovirus ahumpari 3 (HPeV-3), is among the main agents causing severe neonatal neurological infections such as encephalitis and meningitis. However, the underlying molecular mechanisms and changes to the host cellular landscape leading to neurological disease has been understudied. Through quantitative proteomic analysis of HPeV-3 infected neural organoids, we identified unique metabolic changes following HPeV-3 infection that indicate immunometabolic dysregulation. Protein and pathway analyses showed significant alterations in neurotransmission and potentially, neuronal excitotoxicity. Elevated levels of extracellular glutamate, lactate dehydrogenase (LDH), and neurofilament light (NfL) confirmed glutamate excitotoxicity to be a key mechanism contributing to neuronal toxicity in HPeV-3 infection and can lead to apoptosis induced by caspase signaling. These insights are pivotal in delineating the metabolic landscape following severe HPeV-3 CNS infection and may identify potential host targets for therapeutic interventions. | 2024-09-14 | |
IFN-I signaling in type 2 conventional dendritic cells supports TH2 and T follicular helper differentiation after allergen immunization | 10.1101/2024.09.10.612251 | Webb, G. R.; Hilligan, K. L.; Old, S. I.; Tang, S.-C.; Lamiable, O.; Ronchese, F. | Type 2 dendritic cells (DC2s) are essential for TH2 differentiation, but the signaling pathways involved in allergen sensing, DC activation and instruction of CD4+ T cell priming remain unclear. Previous transcriptomic analyses demonstrated a type-I interferon (IFN-I) signature in skin cDC2s following immunization with non-viable larvae of Nippostrongylus brasiliensis (Nb), house dust mite (HDM), and Schistosoma egg antigen (SEA). Blocking IFN-I signaling with anti-IFNAR1 (aIFNAR1) led to reduced TH2 cytokine responses to these antigens, however, the phenotype of cytokine-producing CD4+ T cells was not further defined. Here we show that conditional loss of IFNAR1 signaling in CD11c+ DCs significantly impaired effector TH2 and TFH CD4+ T cell responses to Nb. In vivo proliferation experiments demonstrated reduced numbers of highly divided CD4+ T cells in IFNAR1deltaCD11c mice compared to IFNAR1WT, with the highly divided population comprising both TH2 and TFH. Characterization of the cDC2 compartment by flow cytometry and bulk RNAseq demonstrated lower numbers of Nb+ cDC2s in the skin-draining LN and a reduced expression of Il15 and Il15Ra in IFNAR1dletaCD11c mice compared to IFNAR1WT, while expression of costimulatory molecules including CD80, CD86, Cd40 and Pdcd1lg2 (PD-L2) was not impaired. Therefore, IFN-I conditioning of skin cDC2s is necessary for their effective priming of CD4+ T cell responses to allergens, providing evidence for a role of tissue cytokines in driving cDC2 activation in a TH2 context. | 2024-09-14 | |
High fidelity genetic markers for sexing Cannabis sativa seedlings | 10.1101/2024.09.10.612257 | Prentout, D.; El Aoudati, S.; Mathis, F.; Marais, G.; Henri, H. | The uses of Cannabis sativa, a dioecious species with an XY sex chromosome system, are varying from fiber and oil to cannabinoids, among others. In most cases, males are undesirable and the sexual dimorphism at immature plants is too subtle for reliable phenotypic sexing, making genetic approaches promising. In this technical note, we present a multiplex PCR-mix, that includes two markers of Y-specific coding regions and one autosomal control marker. This PCR-mix, tested across 12 hemp-type cultivars, encompassing approximately 200 individuals, achieved a 99.5% success rate in identifying the sex of C. sativa seedlings. | 2024-09-14 | |
Brownian motion data augmentation: a method to push neural network performance on nanopore sensors | 10.1101/2024.09.10.612270 | Kipen, J.; Jalden, J. | Nanopores are highly sensitive sensors that have achieved commercial success in DNA/RNA sequencing, with potential applications in protein sequencing and biomarker identification. Solid-state nanopores, in particular, face challenges such as instability and low signal-to-noise ratios (SNRs), which lead scientists to adopt data-driven methods for nanopore signal analysis, although data acquisition remains restrictive. In this paper, we augment training samples by simulating virtual Brownian motion based on dynamic models in the literature. We apply this method to a publicly available dataset of a classification task containing nanopore reads of DNA with encoded barcodes. A neural network named QuipuNet was previously published for this dataset, and we demonstrate that our augmentation method produces a noticeable increase in QuipuNets accuracy. Furthermore, we introduce a novel neural network named YupanaNet, which achieves greater accuracy (95.8%) than QuipuNet (94.6%) on the same dataset. YupanaNet benefits from both the enhanced generalization provided by Brownian motion data augmentation and the incorporation of novel architectures, including skip connections and a self-attention mechanism. | 2024-09-14 | |
Identification and Validation of Mitophagy-Related Genes in Diabetic Retinopathy | 10.1101/2024.09.10.612286 | Peng, W.; Zou, Y. | Background: Diabetic retinopathy is one of the common chronic complications of diabetes, characterized by retinal microvascular and neurodegenerative impairment,and it is the primary cause of vision impairment and blindness in adults. Many studies have demonstrated that mitophagy plays a significant role in the pathological mechanism of DR. However, its mechanism is not yet fully clear and requires further research. Methods: We obtained relevant datasets of diabetic retinopathy from the GEO database and used R language to screen for differentially expressed genes. We intersected these genes with mitophagy-related genes and identified differentially expressed mitophagy-related genes. We performed GO and KEGG analysis on the differentially expressed mitophagy-related genes, followed by PPI network analysis. Using Cytoscape software, we selected mitophagy hub genes. Finally, we further validated the expression of the mitophagy hub genes in an in vitro cell culture high-glucose model using quantitative real-time polymerase chain reaction (qRT-PCR). Results: We identified 27 differentially expressed genes related to mitophagy by using R language, with 10 genes upregulated and 17 genes downregulated. We performed GO and KEGG enrichment analysis using R software to further study the potential biological functions of differentially expressed genes. Through PPI network analysis and Cytoscape software, we selected 10 hub genes associated with mitophagy. Finally, through qRT-PCR validation of these 10 hub genes, we found that the mRNA expression differences of MFN1, BNIP3L, GABARAPL1, and PINK1 genes were consistent with our bioinformatics analysis results. Conclusion: We consider that MFN1, BNIP3L, GABARAPL1, and PINK1 may serve as potential biomarkers for diabetic retinopathy.The upregulation and downregulation of these genes provide new insights for further exploration of the role of mitophagy in the pathological mechanism of diabetic retinopathy.These genes can serve as new potential therapeutic targets for the treatment of diabetic retinopathy. | 2024-09-14 | |
Quantum mechanics predicts Bicoid interpretation times of less than a second | 10.1101/2024.09.10.612267 | Lone, I. | The establishment and interpretation of the concentration distribution of the morphogen Bicoid (Bcd) is considered crucial for the successful embryonic development of fruit flies. However, the biophysical mechanisms behind the timely formation and subsequent interpretation of this prototypical morphogenetic system by its target genes are not yet completely understood. Recently a discrete time, one-dimensional quantum walk model of Bcd gradient formation has been successfully used to explain the observed multiple dynamic modes of the Bcd system. However, the question of its precise interpretation by its primary target gene hunchback (hb) remains still unanswered. In this paper it will be shown that the interpretation of the Bcd gradient by its primary target gene hb, with the observed precision of [~] 10%, takes a time period of less than a second, as expected on the basis of recent experimental observations. Furthermore, the quantum walk model is also used to explain certain key observations of recent optogenetic experiments concerning the time windows for Bcd interpretation. Finally, it is concluded that the incorporation of quantum effects into the treatment of Bcd gradient represents a viable step in exploring the dynamics of morphogen gradients. | 2024-09-14 | |
Platelet PI3Kβ regulates breast cancer metastasis | 10.1101/2024.09.10.612261 | Graff, R. C.; Haimowitz, A.; Aguilan, J.; Levine, A.; Zhang, J.; Yuan, W.; Roose-Girma, M.; Seshagiri, S.; Porcelli, S. A.; Gamble, M. J.; Sidoli, S.; Bresnick, A. R.; Backer, J. | Platelets promote tumor metastasis by several mechanisms. Platelet-tumor cell interactions induce the release of platelet cytokines, chemokines, and other factors that promote tumor cell epithelial-mesenchymal transition and invasion, granulocyte recruitment to circulating tumor cells (CTCs), and adhesion of CTCs to the endothelium, assisting in their extravasation at metastatic sites. Previous studies have shown that platelet activation in the context of thrombus formation requires the Class IA PI 3-kinase PI3K{beta}. We now define a role for platelet PI3K{beta} in breast cancer metastasis. Platelet PI3K{beta} is essential for platelet-stimulated tumor cell invasion through Matrigel. Consistent with this finding, in vitro platelet-tumor cell binding and tumor cell-stimulated platelet activation are reduced in platelets isolated from PI3K{beta} mutant mice. RNAseq and proteomic analysis of human breast epithelial cells co-cultured with platelets revealed that platelet PI3K{beta} regulates the expression of EMT and metastasis-associated genes in these cells. The EMT and metastasis-associated proteins PAI-1 and IL-8 were specifically downregulated in co-cultures with PI3K{beta} mutant platelets. PI3K{beta} mutant platelets are impaired in their ability to stimulate YAP and Smad2 signaling in tumor cells, two pathways regulating PAI-1 expression. Finally, we show that mice expressing mutant PI3K{beta} show reduced spontaneous metastasis, and platelets isolated from these mice are less able to stimulate experimental metastasis in WT mice. Taken together, these data support a role for platelet PI3K{beta} in promoting breast cancer metastasis and highlight platelet PI3K{beta} as a potential therapeutic target. | 2024-09-14 | |
Reduction of RAD23A extends lifespan and mitigates pathology in TDP-43 mice | 10.1101/2024.09.10.612226 | Xueshui, G. n.; Prajapati, R. n.; Chun, J. n.; Byun, I. n.; Gebis, K. K.; Wang, Y.-Z. n.; Ling, K. n.; Dalton, C. n.; Blair, J. A.; Hamidianjahromi, A. n.; Bachmann, G. n.; Rigo, F. n.; Jafar-nejad, P. n.; Savas, J. n.; Lee, M. J.; Sreedharan,, J. n.; Kalb, R. | Protein misfolding and aggregation are cardinal features of neurodegenerative disease (NDD) and they contribute to pathophysiology by both loss-of-function (LOF) and gain-of-function (GOF) mechanisms. This is well exemplified by TDP-43 which aggregates and mislocalizes in several NDDs. The depletion of nuclear TDP-43 leads to reduction in its normal function in RNA metabolism and the cytoplasmic accumulation of TDP-43 leads to aberrant protein homeostasis. A modifier screen found that loss of rad23 suppressed TDP-43 pathology in invertebrate and tissue culture models. Here we show in a mouse model of TDP-43 pathology that genetic or antisense oligonucleotide (ASO)-mediated reduction in rad23a confers benefits on survival and behavior, histological hallmarks of disease and reduction of mislocalized and aggregated TDP-43. This results in improved function of the ubiquitin-proteasome system (UPS) and correction of transcriptomic alterations evoked by pathologic TDP-43. RAD23A-dependent remodeling of the insoluble proteome appears to be a key event driving pathology in this model. As TDP-43 pathology is prevalent in both familial and sporadic NDD, targeting RAD23A may have therapeutic potential. | 2024-09-14 | |
Quantitative cytoarchitectural phenotyping of deparaffinized human brain tissues | 10.1101/2024.09.10.612232 | Di Meo, D.; Sorelli, M.; Ramazzotti, J.; Cheli, F.; Bradley, S.; Perego, L.; Lorenzon, B.; Mazzamuto, G.; Emmi, A.; Porzionato, A.; De Caro, R.; Garbelli, R.; Biancheri, D.; Pelorosso, C.; Conti, V.; Guerrini, R.; Pavone, F. S.; Costantini, I. | Advanced 3D imaging techniques and image segmentation and classification methods can profoundly transform biomedical research by offering deep insights into the cytoarchitecture of the human brain in relation to pathological conditions. Here, we propose a comprehensive pipeline for performing 3D imaging and automated quantitative cellular phenotyping on Formalin-Fixed Paraffin-Embedded (FFPE) human brain specimens, a valuable yet underutilized resource. We exploited the versatility of our method by applying it to different human specimens from both adult and pediatric, normal and abnormal brain regions. Quantitative data on neuronal volume, ellipticity, local density, and spatial clustering level were obtained from a machine learning-based analysis of the 3D cytoarchitectural organization of cells identified by different molecular markers in two subjects with malformations of cortical development (MCD). This approach will grant access to a wide range of physiological and pathological paraffin-embedded clinical specimens, allowing for volumetric imaging and quantitative analysis of human brain samples at cellular resolution. Possible genotype-phenotype correlations can be unveiled, providing new insights into the pathogenesis of various brain diseases and enlarging treatment opportunities. | 2024-09-14 | |
Cholinergic feedback for context-specific modulation of sensory representations | 10.1101/2024.09.10.612274 | Yu, B.; Yue, Y.; Ren, C.; Yun, R.; Lim, B.; Komiyama, T. | The brain's ability to prioritize behaviorally relevant sensory information is crucial for adaptive behavior, yet the underlying mechanisms remain unclear. Here, we investigated the role of basal forebrain cholinergic neurons in modulating olfactory bulb (OB) circuits in mice. Calcium imaging of cholinergic feedback axons in OB revealed that their activity is strongly 15 correlated with orofacial movements, with little responses to passively experienced odor stimuli. However, when mice engaged in an odor discrimination task, OB cholinergic axons rapidly shifted their response patterns from movement-correlated activity to odor-aligned responses. Notably, these odor responses during olfactory task engagement were absent in cholinergic axons projecting to the dorsal cortex. The level of odor responses correlated with task 20 performance. Inactivation of OB-projecting cholinergic neurons during task engagement impaired performance and reduced odor responses in OB granule cells. Thus, the cholinergic system dynamically modulates sensory processing in a modality-specific and context-dependent manner, providing a mechanism for a flexible and adaptive sensory prioritization. | 2024-09-14 | |
In the brain of the beholder: bi-stable motion reveals mesoscopic-scale feedback modulation in V1 | 10.1101/2024.09.10.612252 | Pizzuti, A.; Gulban, O. F.; Huber, L.; Peters, J.; Goebel, R. | Understanding the neural processes underlying conscious perception remains a central goal in neuroscience. Visual illusions, whether static or dynamic, provide an effective ecological paradigm for studying conscious perception, as they induce subjective experiences from constant visual inputs. While previous neuroimaging studies have dissociated perceptual interpretation of visual motion from sensory input within the motion-sensitive area (hMT+) in humans, less is known about the role of V1 and its relationship to hMT+ during a bistable perception. To address this, we conducted a layer-fMRI study at 7 T with human participants exposed to a bistable motion quartet stimulus. Despite a constant sensory input, the bistable motion quartet elicits switching horizontal and vertical apparent motion percepts likely due to lateral and feedback connections across low and high-level brain regions (feedback processing). As control, we used an unambiguous version of the motion quartet, hereafter referred to as physical motion stimulus, where horizontal and vertical motion is physically presented as visual stimulus in an alternated fashion (feedforward processing). With the advantage of a sub-millimeter resolution gained at ultra-high field (7 Tesla), we aimed to unveil the differential laminar modulation of V1 (low visual area) and hMT+ (high-visual area) during the physical and bistable condition. Our results indicate that: 1) hMT+ functional activity correlates with conscious perception during both physical and ambiguous stimuli with similar strength. There is no evidence of differential laminar profiles in hMT+ between the two experimental conditions. 2) Between inducer squares, V1 shows a significantly reduced functional response to the ambiguous stimulus compared to the physical stimulus, as it primarily reflects feedback signals with diminished feedforward input. Distinct V1 laminar profiles differentiate the two experimental conditions. 3) The temporal dynamics of V1 and hMT+ become more similar during the ambiguous condition. 4) V1 exhibits reduced specificity to horizontal and vertical motion perception during the ambiguous condition at the retinotopic locations corresponding to the perceived motion. Our findings demonstrate that during the ambiguous condition, there is a stronger temporal coupling between hMT+ and V1 due to feedback signals from hMT+ to V1. Such feedback to V1 might be contributing to the stabilization of the vivid perception of directed motion at the face of constant ambiguous stimulation. | 2024-09-14 | |
Longitudinal trajectories of aperiodic EEG activity in early to middle childhood | 10.1101/2024.09.10.612049 | Sacks, D. D.; Valdes, V.; Wilkinson, C. L.; Levin, A. R.; Nelson, C. A.; Enlow, M. B. | Background: Emerging evidence suggests that aperiodic EEG activity may follow a nonlinear growth trajectory in childhood. However, existing studies are limited by small assessment windows and cross-sectional samples that are unable to fully capture these patterns. The current study aimed to characterize the developmental trajectories of aperiodic activity longitudinally from infancy to middle childhood. We examined potential trajectory differences by sex and brain region. We further investigated whether different developmental trajectories resulted in differential associations between aperiodic activity and maternal anxiety symptoms. Methods: A community sample of children and their parents (N=391) enrolled in a longitudinal study of emotion processing were assessed at infancy, and at ages 3 years, 5 years, and 7 years. Analyses included individual growth curve and mixed effect models. Developmental trajectories of the aperiodic slope and offset were investigated across whole brain, frontal, central, temporal, and posterior regions. Associations of whole brain slope and offset with maternal anxiety symptoms were also examined. Results: Developmental trajectories for both slope and offset were generally characterized by a relative increase in early childhood and a subsequent decrease or stabilization by age 7, with variation by brain region. Females showed relatively steeper slopes at some ages, and males showed relatively greater offset at certain ages. Maternal anxiety was negatively associated with slope at 3 years and positively associated with slope at 7 years. Conclusions: The longitudinal developmental trajectory of aperiodic slope in early childhood is nonlinear and shows variation by sex and brain region. The magnitude and direction of associations with maternal anxiety varied by age, corresponding with changes in trajectories. Developmental stage should be considered when interpreting findings related to aperiodic activity in childhood. | 2024-09-14 | |
G-CSF and IL-6 drive myeloid dysregulation during severe viral infection | 10.1101/2024.09.12.612764 | Kajihara, K.; Yan, D.; Seim, G.; Little-Hooy, H.; Kang, J.; Chen, C.; De Simone, M.; Delemarre, T.; Darmanis, S.; Shivram, H.; Bauer, R.; Rosenberger, C. M.; Kapadia, S.; Xu, M.; Reyes, M. | Dysregulated myeloid states are associated with disease severity in both sepsis and COVID-19. However, their relevance in non-COVID-19 viral infection, the factors driving their induction, and their role in tissue injury remain poorly understood. We performed a meta-analysis of 1,622,180 myeloid cells from 890 COVID-19 or sepsis patients and controls across 19 published blood scRNA-seq datasets, which revealed severity-associated gene programs in both neutrophils and monocytes pointing to emergency myelopoiesis (EM). Using published bulk transcriptional data from 562 individuals with non-COVID-19 viral disease, we show that these signatures are similarly upregulated during severe influenza and RSV infection. Analysis of transcriptional and proteomic responses in tocilizumab-treated COVID-19 patients show that IL-6 signaling blockade results in a partial reduction of EM signatures and a compensatory increase in the growth factor G-CSF. Using a cellular model of human myelopoiesis, we show that both IL-6 and G-CSF stimulate the production of myeloid cells that express EM signatures in vitro. Using a mouse model of severe influenza infection, we demonstrate the effect of IL-6 and G-CSF signaling blockade on EM-associated myeloid cells, and highlight the opposing effects of EM-induced neutrophils and monocytes on tissue injury. Our study demonstrates the link between systemic cytokines and myeloid dysregulation during severe infection in humans, and highlights the cooperative role of IL-6 and G-CSF signaling in driving infection-induced myelopoiesis. | 2024-09-14 | |
BRUCE liver-deficiency potentiates MASLD/MASH in PTEN liver-deficient background by impairment of mitochondrial metabolism in hepatocytes and activation of STAT3 signaling in hepatic stellate cells | 10.1101/2024.09.13.611500 | Che, L.; Stevenson, C. K.; Plas, D. R.; Wang, J.; Du, C. | Metabolic dysfunction-associated steatotic liver disease (MASLD) is currently the most common liver disease, affecting up to 25% of people worldwide, featuring excessive fat accumulation in hepatocytes. Its advanced form, metabolic dysfunction-associated steatohepatitis (MASH), is a serious disease with hepatic inflammation and fibrosis, increasing the need for liver transplants. However, the pathogenic mechanism of MASLD and MASH is not fully understood. We reported that BRUCE (BIRC6) is a liver cancer suppressor and is downregulated in MASLD/MASH patient liver specimens, though the functional role of BRUCE in MASLD/MASH remains to be elucidated. To this end, we generated liver-specific double KO (DKO) mice of BRUCE and PTEN, a major tumor suppressor and MASLD/MASH suppressor. By comparing liver histopathology among 2-3-month-old mice, there were no signs of MASLD or MASH in BRUCE liver-KO mice and only onset of steatosis in PTEN liver-KO mice. Interestingly, DKO mice had developed robust hepatic steatosis with inflammation and fibrosis. Further analysis of mitochondrial function with primary hepatocytes found moderate reduction of mitochondrial respiration, ATP production and fatty acid oxidation in BRUCE KO and the greatest reduction in DKO hepatocytes. Moreover, aberrant activation of pro-fibrotic STAT3 signaling was found in hepatic stellate cells (HSCs) in DKO mice which was prevented by administered STAT3-specific inhibitor (TTI-101). Collectively, the data demonstrates by maintaining mitochondrial metabolism BRUCE works in concert with PTEN to suppress the pro-fibrogenic STAT3 activation in HSCs and consequentially prevent MASLD/MASH. The findings highlight BRUCE being a new co-suppressor of MASLD/MASH. | 2024-09-14 | |
Ongoing evolution of Middle East Respiratory Syndrome Coronavirus, Kingdom of Saudi Arabia, 2023-2024 | 10.1101/2024.09.12.612455 | Hassan, A. M.; Mühlemann, B.; Al-Subhi, T. L.; Rodon, J.; El-Kafrawy, S. A.; Memish, Z.; Melchert, J.; Bleicker, T.; Mauno, T.; Perlman, S.; Zumla, A.; Jones, T. C.; Müller, M. A.; Corman, V. M.; Drosten, C.; Azhar, E. I. | Middle East respiratory syndrome coronavirus (MERS-CoV) circulates in dromedary camels in the Arabian Peninsula and occasionally causes spillover infections in humans. Due to lack of sampling during the SARS-CoV-2 pandemic, current MERS-CoV diversity is poorly understood. Of 558 dromedary camel nasal swabs from Saudi Arabia, sampled November 2023 to January 2024, 39% were positive for MERS-CoV RNA by RT-PCR. We generated 42 MERS-CoV and seven human 229E-related CoV by high-throughput sequencing. For both viruses, the sequences fell into monophyletic clades apical to the most recent available genomes. The MERS-CoV sequences were most similar to those from lineage B5. The new MERS-CoVs sequences harbor unique genetic features, including novel amino acid polymorphisms in the Spike protein. The new variants require further phenotypic characterization to understand their impact. Ongoing MERS-CoV spillovers into humans pose significant public health concerns, emphasizing the need for continued surveillance and phenotypic studies. | 2024-09-14 | |
SCAMPI: A scalable statistical framework for genome-wide interaction testing harnessing cross-trait correlations | 10.1101/2024.09.10.612314 | Bian, S.; Bass, A. J.; Liu, Y.; Wingo, A. P.; Wingo, T.; Cutler, D. J.; Epstein, M. P. | Family-based heritability estimates of complex traits are often considerably larger than their single-nucleotide polymorphism (SNP) heritability estimates. This discrepancy may be due to non-additive effects of genetic variation, including variation that interacts with other genes or environmental factors to influence the trait. Variance-based procedures provide a computationally efficient strategy to screen for SNPs with potential interaction effects without requiring the specification of the interacting variable. While valuable, such variance-based tests consider only a single trait and ignore likely pleiotropy among related traits that, if present, could improve power to detect such interaction effects. To fill this gap, we propose SCAMPI (Scalable Cauchy Aggregate test using Multiple Phenotypes to test Interactions), which screens for variants with interaction effects across multiple traits. SCAMPI is motivated by the observation that SNPs with pleiotropic interaction effects induce genotypic differences in the patterns of correlation among traits. By studying such patterns across genotype categories among multiple traits, we show that SCAMPI has improved performance over traditional univariate variance-based methods. Like those traditional variance-based tests, SCAMPI permits the screening of interaction effects without requiring the specification of the interaction variable and is further computationally scalable to biobank data. We employed SCAMPI to screen for interacting SNPs associated with four lipid-related traits in the UK Biobank and identified multiple gene regions missed by existing univariate variance-based tests. SCAMPI is implemented in software for public use. | 2024-09-14 | |
Subcellular Level Spatial Transcriptomics with PHOTON | 10.1101/2024.09.10.612328 | Rajachandran, S.; Xu, Q.; Cao, Q.; Zhang, X.; Chen, F.; Mangiameli, S. M.; Chen, H. | The subcellular localization of RNA is closely linked to its function. Many RNA species are partitioned into organelles and other subcellular compartments for storage, processing, translation, or degradation. Thus, capturing the subcellular spatial distribution of RNA would directly contribute to the understanding of RNA functions and regulation. Here, we present PHOTON (Photoselection of Transcriptome over Nanoscale), a method which combines high resolution imaging with high throughput sequencing to achieve spatial transcriptome profiling at subcellular resolution. We demonstrate PHOTON as a versatile tool to accurately capture the transcriptome of target cell types in situ at the tissue level such as granulosa cells in the ovary, as well as RNA content within subcellular compartments such as the nucleolus and the stress granule. Using PHOTON, we also reveal the functional role of m6A modification on mRNA partitioning into stress granules. These results collectively demonstrate that PHOTON is a flexible and generalizable platform for understanding subcellular molecular dynamics through the transcriptomic lens. | 2024-09-14 | |
Exploring American Elderberry Compounds for Antioxidant, Antiviral, and Antibacterial Properties Through High-Throughput Screening Assays Combined with Untargeted Metabolomics | 10.1101/2024.09.13.611920 | Dwikarina, A.; Bayati, M.; Efrat, N.; Roy, A.; Lei, Z.; Ho, K.-V.; Sumner, L. W.; Thomas, A. L.; Applequist, W.; Michael Greenlief, M.; Townesmith, A.; Lin, C.-H. | American elderberry (Sambucus nigra subsp. canadensis) is a rapidly emerging new perennial crop for Missouri, recognized for its high level of bioactive compounds with significant health benefits, including antimicrobial, antiviral, and antioxidant properties. A high-throughput screening assay combined with untargeted metabolomics analysis was utilized on American elderberry juice from 21 genotypes to explore and characterize these bioactive compounds. Our metabolomics study has identified 32 putative bioactive compounds in the American Elderberry juices. An array of high-throughput screening bioassays evaluated 1) total antioxidant capacity, 2) activation of antioxidant response elements (ARE), 3) antiviral activity, and 4) antibacterial activity of the putatively identified compounds. Our results revealed that 14 of the 32 American elderberry compounds exhibited strong antioxidant activity. Four compounds (isorhamnetin 3-O-glucoside, kaempferol, quercetin, and naringenin) activated ARE activity and were found to be non-cytotoxic to cells. Notably, six of the 32 compounds demonstrated significant antiviral activity in an in vitro TZM-bl assay against two strains of HIV-1 virus, CXCR4-dependent NL4-3 virus and CCR5-dependent BaL virus. Luteolin showed the most potent anti-HIV activity in an in vitro TZM-bl assay against the NL4-3 virus (IC50 = 1.49 microM), followed by isorhamnetin (IC50 = 1.67 microM). The most potent anti-HIV compound against the BaL virus was myricetin (IC50 = 1.14 microM), followed by luteolin (IC50 = 4.38 microM). Additionally, six compounds were found to have antibacterial activity against gram-positive bacteria S. aureus, with cyanidin 3-O-rutinoside having the most potent antibacterial activity in vitro (IC50 = 2.9 microM), followed by cyanidin 3-O-glucoside (IC50 = 3.7 microM). These findings support and validate the potential health benefits of compounds found in American elderberry juices, and highlight their potential for use in dietary supplements as well as innovative applications in health and medicine. | 2024-09-14 | |
Rapid production of modified cattle and sheep from spermatid-like stem cells | 10.1101/2024.09.10.612264 | Yang, L.; Di, A.; Song, L.; Liu, X.; Wu, D.; Wang, S.; Hao, Z.; Bu, L.; Bai, C.; Su, G.; Wei, Z.; Zhang, L.; Liu, Z.; Gao, S.; Li, G. | Haploid androgenetic stem cells (haSCs) are revolutionary resources for studying animal traits and greatly impact livestock breeding, but livestock haSCs have yet to be obtained. Here, we report the derivation of cattle and sheep haSCs using a novel FACE medium, and these cells harbor formative pluripotency features. Notably, knockout cattle and sheep can be efficiently obtained upon injection of the protaminized haSCs with targeted gene mutations into oocytes. | 2024-09-14 | |
Randomly oriented microgrooved hydrogel guides cellular motility, modulates speed, and governs directionality of cellular spread | 10.1101/2024.09.10.612339 | Ghosh, B.; Agarwal, K. | Cell migration is a fundamental biological process, yet the mechanisms underlying how cells sense and navigate complex environments remain poorly understood. In this study, we developed a system of randomly oriented microgrooves, designed at cellular length scales, to explore motility intelligence in response to varied topographies. These microgrooves allowed cells to freely choose their migratory paths, revealing key insights into how cells sense and adapt to topological cues. Using fibroblast cells migrating over these grooved substrates, we examined cellular processes such as actin cytoskeleton remodeling, cell adhesion dynamics, and the impact of groove alignment on migration speed and directionality. Our results demonstrate that cells align their cytoskeletal structures to groove geometries, forming actin-rich anchors that enhance migration in groove-aligned environments. Cells migrating in grooves aligned with their intrinsic polarity exhibited faster, more directed migration compared to those in misaligned or control conditions. This work advances our understanding of cell-topology interaction and provides new perspectives for tissue engineering applications in cancer therapy and wound healing. | 2024-09-14 | |
Enantiomeric histidine-rich peptide coacervates enhance antigen delivery to T cells | 10.1101/2024.09.10.612317 | Pramanik, U.; Das, A.; Brown, E. M.; Struckman, H. L.; Wang, H.; Stealey, S.; Sprunger, M. L.; Abdul, W.; Fascetti, J.; Mondal, J.; Silva, J. R.; Zustiak, S. P.; Jackrel, M. E.; Rudra, J. S. | Peptides and peptidomimetics that self-assemble via LLPS have recently emerged as building blocks for fabricating functional biomaterials due to their unique physicochemical properties and dynamic nature. One of life's most distinctive signatures is its selectivity for chiral molecules and, to date, coacervates comprised of D-amino acids have not been reported. Here, we demonstrate that histidine-rich repeats of (GHGXY)4 (X=L/V/P) and their enantiomers undergo LLPS opening new avenues for enhancing coacervate stability. Through a series of biophysical studies, we find that LLPS kinetics, droplet size, fusion, and encapsulation efficiency are dictated by the primary sequence. Further, these coacervates can encapsulate therapeutic cargo which are then internalized via endocytic mechanisms. Finally, we show that the coacervates enhance antigen presentation to CD4+ and CD8+ T cells resulting in robust proliferation and production of functional cytokines. Collectively, our study describes the development and characterization of enantiomeric peptide coacervates as attractive vaccine delivery vehicles with tunable physicochemical properties. | 2024-09-14 | |
Associations on the Fly, a new feature aiming to facilitate exploration of the Open Targets Platform evidence | 10.1101/2024.09.10.612089 | Cruz-Castillo, C.; Fumis, L.; Mehta, C.; Martinez-Osorio, R. E.; Roldan-Romero, J. M.; Cornu, H.; Uniyal, P.; Solano-Roman, A.; Carmona, M.; Ochoa, D.; McDonagh, E. M.; Buniello, A. | The Open Targets Platform (https://platform.opentargets.org) is a unique, comprehensive, open-source resource supporting systematic identification and prioritisation of targets for drug discovery. The Platform combines, harmonises and integrates data from >20 diverse sources to provide target-disease associations, covering evidence derived from genetic associations, somatic mutations, known drugs, differential expression, animal models, pathways and systems biology. An in-house target identification scoring framework weighs the evidence from each data source and type, contributing to an overall score for each of the 7.8M target-disease associations. However, the previous infrastructure did not allow user-led dynamic adjustments in the contribution of different evidence types for target prioritisation, a limitation frequently raised by our user community. Furthermore, the previous Platform user interface did not support navigation and exploration of the underlying target-disease evidence on the same page, occasionally making the user journey counterintuitive. Here, we describe Associations on the Fly (AOTF), a new Platform feature - developed as part of a wider product refactoring project - to enable formulation of more flexible and impactful therapeutic hypotheses through dynamic adjustment of the weight of contributing evidence from each source, altering the prioritisation of targets. | 2024-09-14 | |
3D Molecular Pretraining via Localized Geometric Generation | 10.1101/2024.09.10.612249 | Sun, Y.; Chen, K.; Liu, K.; Ye, Q. | Self-supervised learning on 3D molecular structures is gaining importance in data-driven scientific research and applications due to the high costs of annotating biochemical data. However, the strategic selection of semantic units for modeling 3D molecular structures remains underexplored, despite its crucial role in effective pretraining - a concept well-established in language processing and computer vision. We introduce Localized Geometric Generation (LEGO), a novel approach that treats tetrahedrons within 3D molecular structures as fundamental building blocks, leveraging their geometric simplicity and widespread presence across chemical functional patterns. Inspired by masked modeling, LEGO perturbs tetrahedral local structures and learns to reconstruct them in a self-supervised manner. Experimental results demonstrate LEGO consistently enhances molecular representations across biochemistry and quantum property prediction benchmarks. Additionally, the tetrahedral modeling and pretraining generalize from small molecules to larger molecular systems, validating by protein-ligand affinity prediction. Our results highlight the potential of selecting semantic units to build more expressive and interpretable neural networks for scientific AI applications. | 2024-09-14 | |
Distinct Stromal Cell Populations Define the B-cell Acute Lymphoblastic Leukemia Microenvironment | 10.1101/2024.09.10.612346 | Blanco, M. N. F.; Kazybay, B.; Belderbos, M.; Heidenreich, O.; Vormoor, J. H. | The bone marrow microenvironment is critical for B-cell acute lymphoblastic leukemia (B-ALL) but its cellular heterogeneity remains poorly defined. Here, we employed single-cell RNA sequencing to comprehensively characterize the stromal and hematopoietic niches in pediatric B-ALL. Our analysis revealed two distinct mesenchymal stromal cell (MSC) populations as primary leukemia-supportive niches: early mesenchymal progenitors and adipogenic progenitors. Single-cell transcriptomic analysis infers that ALL blasts use distinct cell-cell interactions to communicate with the different stromal populations. Purified adipogenic progenitors from the bone of children with ALL support survival of the leukemic blasts ex vivo and their signature is enriched in relapse samples. Our data establish adipogenic progenitors as a distinct and novel component of the ALL niche. | 2024-09-14 | |
Optimizing The Fabrication Of Shape-Defined Microparticles For Sustained Drug Delivery: The 'Less Is More' Paradigm | 10.1101/2024.09.10.612107 | Murgia, D.; Martins Estevao, B.; Portioli, C.; Palomba, R.; Decuzzi, P. | Polymeric microparticles find extensive use in several pharmaceutical applications. Our group has developed poly(lactic-co-glycolic acid) (PLGA) microPLates (PL) featuring a square base of 20 x 20 m and a height of 10 m for the controlled and sustained delivery of a range of therapeutic payloads, including anti-inflammatory and anti-cancer drugs, small molecules for neurodevelopmental disorders, and siRNA for osteoarthritis. In this study, the morphological and pharmacological properties of PLGA-PL were optimized by introducing new steps in the original fabrication protocol and systematically varying the polymer content. Vacuum suction was used to control solvent removal, and two different "cleaning" steps were tested, resulting in six different PL configurations with a PLGA content ranging from 2 to 10 mg. Electron and optical microscopy analyses confirmed the well-defined square shape of PL, with a central concavity depending on the PLGA content. Fabrication yielding ranged between 10% and 70%, while encapsulation efficiencies reached approximately 15% using curcumin (CURC) as a model drug. The kinetics of CURC release was analyzed using the semi-empirical model of Korsmeyer-Peppas, suggesting either a Fickian diffusion or anomalous transport mechanisms based on the PLGA amounts. Complementary techniques were used to assess morphological alterations and mass loss, evaluating the degradation PL over time in water and physiological solutions. Unexpectedly, PL configurations with lower PLGA contents exhibited higher fabrication yielding, drug encapsulation, and slower drug release. The optimized fabrication approach offers greater flexibility to tailor the degradation and pharmacological properties of PL for various therapeutic applications. | 2024-09-14 | |
The preferential injury of outer renal medulla after ischemia-reperfusion relies on high oxidative metabolism | 10.1101/2024.09.12.612245 | Arnoux, G.; Serre, J.; Verissimo, T.; Tihy, M.; Placier, S.; Verney, C.; Sangla, F.; Paolucci, D.; Fernandez, M.; de Seigneux, S.; Sgardello, S.; Naesens, M.; Hadchouel, J.; Feraille, E.; Khodo, S. N.; Galichon, P.; Legouis, D. | Acute kidney injury (AKI) is a prevalent and significant complication in critically ill patients, and its management remains a considerable challenge. The kidney is a highly metabolic organ, consuming and producing substantial amounts of ATP, mainly through mitochondrial oxidative phosphorylation. Recently, mitochondrial dysfunction has been identified as a key factor in the pathophysiology of AKI and the progression to chronic kidney disease. The kidney is a complex organ, comprising millions of structural and functional units. These nephrons are composed of different cell types dwelling within specific metabolic microenvironment. Whether the metabolic spatialization in the kidney has consequences on tubular injury distribution and severity remains unclear. In this study, we identified the high metabolic rate of the outer stripe of the outer medulla (OSOM) and its substrate preference flexibility, relying on both glycolysis and fatty acid oxidation (FAO) to fulfill its ATP demands. We demonstrated that the OSOM is susceptible to mitochondrial and FAO impairment induced by propofol, the most used sedative in intensive care settings, which exacerbates tubular injury during AKI. In the clinical setting, the cumulative dose of propofol is positively correlated with oxidative metabolism disruption and histological and function outcomes in renal allograft recipients. Finally, we found that the loop of Henle, the OSOM major constituent, was the most injured segment during AKI in patients. This study shows how renal metabolic spatialization impacts tubular injury severity. We identified the OSOM as the most metabolically active and the most injured region of the kidney both in humans and mice. We demonstrated that propofol is a potent inhibitor of renal mitochondrial function and FAO exacerbating tubular injury in the OSOM upon IRI. | 2024-09-14 | |
Proof of Concept for High-Dose Cannabidiol Pretreatment to Antagonize Opioid Induced Persistent Apnea | 10.1101/2024.09.13.612358 | Wiese, B. M.; Bondarenko, E.; Feldman, J. L. | Using a mouse equivalent of FDA-approved cannabidiol (CBD) dosing, we found high dose CBD affects opioid induced persistent apnea (OIPA), the principal cause of opioid related fatalities. CBD pretreatment mitigated respiratory depression from fentanyl in awake mice and significantly delayed OIPA onset in anesthetized mice, effective as the opioid antagonist naloxone. The powerful effect of CBD pretreatment on OIPA suggests a novel therapeutic strategy to reduce fatal opioid overdose incidence. | 2024-09-14 | |
Distinct roles of general anesthesia activated CeA neurons in acute versus late phase of neuropathic pain | 10.1101/2024.09.11.612553 | Zhao, J.; Furutani, K.; McGinnis, A.; Mathew, J. P.; Wang, F.; Ji, R. R. | A previous study discovered a distinct population of GABAergic neurons in the central amygdala (CeA) that can be activated by general anesthesia (CeA-GA) and exert analgesic functions (Hua et al., 2020). To independently reproduce these prior findings and to investigate the electrophysiological properties of CeA-GA neurons, we first used 1.2 percent isoflurane to induce c-Fos activation in the mouse brain and validated the Fos expression by RNAscope in situ hybridization. Indeed, isoflurane induced robust Fos expression in CeA and these Fos+ CeA-GA neurons are GABAergic neurons (Vgat+). We next used Fos-TRAP2 method (different from the CANE method used in the prior study) to label CeA-GA neurons (tdTomato+). Our ex vivo electrophysiological recordings in brain slices revealed that compared to Fos-negative CeA neurons, CeA-GA neurons had significantly higher excitability and exhibited distinct patterns of action potentials. Chemogenetic activation of Fos-TRAPed CeA-GA neurons was effective at increasing pain thresholds in naive mice and mice with early-phase neuropathic pain 2 weeks after spared nerve injury (SNI). However, the same chemogenetic activation of CeA-GA neurons only had modest analgesia in the late phase of SNI at 8 weeks, although it was highly effective in reducing chronic pain-associated anxiety behaviors at this stage. We found that Fos-negative CeA neurons, but not CeA-GA neurons, exhibited increased excitability in the late-phase of SNI, suggesting that chronic pain causes a shift in the relative activity of the CeA microcircuit. Interestingly, Fos-negative neurons exhibited much higher expression of K/Cl cotransporter-2 (KCC2), and KCC2 expression was downregulated in the CeA in the late-phase of neuropathic pain. These results support the idea that targeting CeA-GA neurons may provide therapeutic benefits for pain relief and chronic pain-associated anxiety. Our findings also suggest distinct roles of CeA-GA neurons in regulating physiological pain, acute pain, and chronic pain with a possible involvement of KCC2. | 2024-09-14 | |
Anisotropy of object non-rigidity: High-level perceptual consequences of cortical anisotropy | 10.1101/2024.09.10.612333 | Maruya, A.; Zaidi, Q. | This study investigates a compelling instance where variations in a complex, higher-level perceptual phenomenon. Specifically, the anisotropy in object non-rigidity are explained by the distribution of low-level neural properties in the primary visual cortex. Through a combination of mathematical derivations and computational simulations, we replicate psychophysical experiment results that explore the perception of non-rigidity in what should be rigidly rotating objects. Specifically, we examine the visual interpretation of two connected circular rings in rotation. When these rigidly connected rings rotate horizontally, observers predominantly perceive rigid rotation. However, when the image is rotated by 90 degrees, observers perceive only non-rigid wobbling. Additionally, vertically elongated rings appear narrower and longer compared to their horizontal counterparts. Our analysis first shows that these perceived shape changes can be decoded from V1 outputs by considering anisotropies in orientation-selective cells. We then empirically demonstrate that even when the vertically rotating ellipses are widened or the horizontally rotating ellipses are elongated to match shapes, the perceived difference in non-rigidity is reduced, but increased non-rigidity persists in vertical rotations, suggesting that the motion mechanism also plays role. By incorporating cortical anisotropies into motion flow computations, we found that the motion gradients for vertical rotations more closely align with physical wobbling, while horizontal rotations fall between wobbling and rigid rotation. This suggests that intrinsic cortical anisotropies contribute to the heightened perception of non-rigidity when orientation shifts from horizontal to vertical. The study underscores the importance of these cortical anisotropies in shaping perceptual outcomes and raises intriguing questions about their evolutionary significance, particularly regarding their impact on shape constancy and motion perception. | 2024-09-14 | |
Pathogenic microbiota disrupts the intact structure of cerebral organoids by altering energy metabolism | 10.1101/2024.09.11.612577 | Isik, M.; Eylem, C. C.; Erdogan-Gover, K.; Aytar-Celik, P.; Enuh, B. M.; Emregul, E.; Cabuk, A.; Yildirim, Y.; Nemutlu, E.; Muotri, A. R.; Derkus, B. | This study investigated the impact of different bacterial populations on the biomolecular structures of cerebral organoids (COs) at various levels. COs were co-cultured with non- pathogenic (NM) and pathogenic (PM) bacterial populations. PM reduced the number of TUJ1+ neurons and disrupted the intact structure of COs. In addition, PM was found to induce changes in the transcript profile of COs, including a decrease in the activity of the glycolysis pathway and an increase in the pentose phosphate pathway, leading to deterioration in cellular energy metabolism, which is linked to neurodegenerative diseases. Proteomic analysis revealed a unique cluster of proteins in COs. PM exposure upregulated proteins related to neurological diseases, consistent with RNA-seq data. Communication between bacteria and neural cells was demonstrated using 18O-stable isotope labeling (SIL)-based metabolic flux analysis. COs showed higher 18O-enrichment of TCA cycle intermediates when co-cultured with NM and PM, indicating increased oxidative phosphorylation activity upon exposure to bacteria. This study provides a useful platform to monitor metabolic signals and communication between microbiotas and human brain cells. The findings suggest that pathogenic bacteria release metabolites that alter biomolecular structures in brain organoids, potentially contributing to neurodegenerative diseases. | 2024-09-14 | |
Influence of the Glymphatic System on α-Synuclein Propagation: Role of Aquaporin-4 and the Dystrophin-Associated Protein Complex | 10.1101/2024.09.11.612272 | Lopes, D. M.; Llewellyn, S. K.; Bury, S. E.; Wang, J.; Wells, J. A.; Gegg, M.; Verona, G.; Lythgoe, M. F.; Harrison, I. F. | Propagation and aggregation of prion proteins, such as tau and -synuclein (Syn), are key pathological features of neurodegenerative diseases. Extracellular clearance pathways, such as the glymphatic system, may play a crucial role in the removal of these toxic proteins from the brain. Primarily active during sleep, this system relies on aquaporin-4 (AQP4) water channel expression and polarisation to astrocytic endfeet, facilitating interstitial solute clearance. Glymphatic dysfunction has recently been implicated in Parkinsons disease, however the precise mechanisms underlying the pathogenic effect of this dysfunction remain unclear. This includes how impaired glymphatic function influences Syn propagation dynamics, and the role of propagating Syn itself on glymphatic function. In this study, we used a mouse model of Syn propagation to elucidate the impact of Syn aggregation on glymphatic function, by measuring CSF-ISF exchange and assessing AQP4 and associated endfoot complex proteins in the brain over time and across different regions. Our results show that direct injection of Syn pre-formed fibrils leads to reduced expression of the AQP4 endfoot complex, but propagation of endogenous Syn induces an enhancement of glymphatic function suggesting compensatory upregulation in response to increasing endogenous Syn burden. To determine the influence of glymphatic dysfunction on Syn propagation dynamics, we then employed a pharmacological approach to inhibit glymphatic function in this model. Acute glymphatic inhibition significantly reduced brain to CSF Syn clearance, and chronic treatment exacerbated Syn pathology, neurodegeneration, and motor behavioural deficits in mice. Together our findings show that Syn clearance and propagation are modulated by glymphatic function and suggest that AQP4 complex dysregulation may contribute to glymphatic impairment associated with Parkinsons diseases. | 2024-09-14 | |
Validity of a New Stress Induction Protocol using Speech Improvisation (IMPRO) | 10.1101/2024.09.10.612289 | Saskovets, M.; Lohachov, M.; Liang, Z.; Piumarta, I. | This paper proposes a novel stress induction protocol utilizing speech improvisation -- Interactive Multitask Performance Response Observation (IMPRO) -- in front of an audience. This approach aims to induce stress through a combination of public speaking, cognitive effort, and challenge to maintain a spontaneous narrative. We investigate the validity of this novel approach by examining physiological responses of 35 healthy participants aged 18 to 38. Saliva cortisol was measured as the ground truth for mental stress assessment. In addition, electrodermal activity (EDA) was used as a non-invasive measure of sympathetic activation, offering real-time data with high resolution. Functional near-infrared spectroscopy (fNIRS) measurement was performed to assess the cortical hemodynamic responses induced by the IMPRO protocol. We focused on hemodynamics in the prefrontal cortex (PFC), a region associated with stress processing. We found that cortisol levels and EDA significantly increased in response to the stress task compared to the baseline. We also observed a significant change in PFC hemodynamic levels in a set of channels compared to the baseline, with a higher overall increase in the right frontopolar area compared to the left. In conclusion, our findings validated IMPRO as an effective and easy to perform method for mental stress induction. | 2024-09-14 | |
Operant conditioning of cortical waves through a brain-machine interface | 10.1101/2024.09.10.612344 | Dogadov, A.; Shulz, D. E.; Ferezou, I.; Ego-Stengel, V.; Estebanez, L. | At the surface of the cerebral cortex, activity dynamics measured at a mesoscopic (about 0.1 to 1 mm in mouse cortex) scale are characterized by both spontaneous and behavior-related dynamical waves of synchronized neuronal activity. These waves are thought to participate in information propagation and processing, but remain poorly understood. To assess if such mesoscopic coordinated neuronal dynamics can be controlled in a goal-directed manner, we implemented a task in which mice are trained to generate waves with specific trajectories in order to obtain rewards. We tracked propagating waves at the surface of the somatosensory-motor cortex of head-fixed mice in real time by means of wide field calcium imaging, and conditioned the delivery of rewards to the detection of wave trajectories responding to specific spatiotemporal criteria. We found that the majority of the trained mice significantly increased their performance, mainly by increasing the frequency of rewardable waves. As the mice learned to achieve this task, we observed changes in the spatiotemporal patterns of the cortical waves. By revealing that, upon learning, neuronal activity can be shaped at the mesoscopic scale to generate specific waves patterns, our work opens up new perspectives for brain-machine interfacing. | 2024-09-14 | |
Protective variant in PLCgamma2 mitigates Alzheimer's disease associated pathology via enhancing beneficial microglia functions | 10.1101/2024.09.12.612207 | Takalo, M.; Jeskanen, H.; Rolova, T.; Kervinen, I.; Hellen, M.; Heikkinen, S.; Koivisto, H.; Jokivarsi, K.; Muller, S. A.; Koivumaki, E.-M.; Makinen, P.; Juopperi, S.-P.; Willman, R.-M.; Sinisalo, R.; Hoffman, D.; Jantti, H.; Peitz, M.; Fliessbach, K.; Kuulasmaa, T.; Natunen, T.; Kemppainen, S.; Poutiainen, P.; Leinonen, V.; Malm, T.; Martiskainen, H.; Ramirez, A.; Haapasalo, A.; Lichtenthaler, S. F.; Tanila, H.; Haass, C.; Rinne, J.; Koistinaho, J.; Hiltunen, M. | PLCgamma2-P522R (phospholipase C gamma 2, proline 522 to arginine) is a protective variant that reduces the risk for late onset Alzheimer's disease (LOAD). Recently, it was shown to decrease beta-amyloid pathology in 5XFAD mouse model of AD. In this study, our goal was to investigate the protective functions of PLCgamma2-P522R variant in less aggressive mouse model of AD as well as to assess underlying mechanisms at the molecular and cellular level using mouse and human microglia models. The effects of the protective PLCgamma2-P522R variant on microglia activation, AD-related beta-amyloid and neuronal pathologies, as well as behavioral changes were investigated in PLCgamma2-P522R knock-in mice crossbred with an APP/PS1 mouse model of AD. Transcriptomic, proteomic, and functional studies were carried out in cultured and acutely isolated adult PLCgamma2-P522R mouse microglia to study molecular mechanisms. Finally, microglia-like cell models generated from blood and skin biopsy samples of the PLCgamma2-P522R variant carriers were employed to translate the key findings in human cells. Our results demonstrate that the PLCgamma2-P522R variant reduces brain beta-amyloid plaque burden of APP/PS1 mice. Simultaneously, PLCgamma2-P522R variant increased non-proinflammatory microglia activation and microglia clustering around beta-amyloid plaques, leading to reduced beta-amyloid plaque-associated neuronal dystrophy. In cultured mouse primary microglia, PLCgamma2-P522R variant decreased accumulation of large lipid droplets, reduced cell stress, and increased acute response to strong inflammatory stimuli. Transcriptomic and proteomic analyses in acutely isolated adult mouse microglia as well as in human monocyte-derived microglial cells showed that PLCgamma2-P522R upregulates mitochondrial fatty acid oxidation and downregulates inflammatory/interferon signaling pathways. Accordingly, PLCgamma2-P522R increased mitochondrial respiration in iPSC -derived microglial cells. Together, these findings suggest that PLCgamma2-P522R variant exerts protection against AD-associated beta-amyloid and neuronal pathologies via enhancing microglial barrier formation around beta-amyloid plaques, but suppressing pro-inflammatory activation. Observed changes in fatty acid metabolism and mitochondrial flexibility as well as the downregulation of genes involved in inflammatory signaling pathways suggest that these protective effects of the PLCgamma2-P522R variant are mediated through an anti-ageing mechanism. | 2024-09-14 | |
Adult oligodendrogenesis gates arcuate neuronal glucose sensing through remodelling of the blood-hypothalamus barrier via ADAMTS4 | 10.1101/2024.09.10.612227 | Buller, S.; Staricoff, E.; Riches, C.; Tsang, A.; Josipovic, M.; Ikemura, K.; Opoku, G.; Sato, I.; Hirohata, S.; Stenzel, S.; Nayar, S.; Vega, M. R.; Hecksher-Sorensen, J.; Timmler, S.; Dowsett, G.; Lam, B.; Yeo, G. S. H.; Alonge, K. M.; Li, H.; Richardson, W. D.; Evans, M. L.; Blouet, C. | Brain glucose sensing is critical for survival during hypoglycaemia and tunes the level of defended blood glucose, which goes up in diabetes. Neuronal glucose sensing neurons and mechanisms have been identified, but how these neurons access blood concentrations of glucose to adjust their output and maintain glucose homeostasis is unclear. Here, we demonstrate that adult oligodendrogenesis in the median eminence (ME) is modulated by changes in circulating glucose levels and rapidly upregulated by hypoglycaemia. We show that genetic blockade of new OL production in adult mice impairs the regulation of glucose homeostasis, the integrity of the ME blood-hypothalamus barrier, and neuronal glucose sensing in the arcuate nucleus of the hypothalamus (ARH). Unexpectedly, functional integrity of adult-formed myelin is not required for the maintenance of glucose homeostasis. Instead, newly formed OLs exert their glucoregulatory actions via the synthesis of A disintegrin and metallopeptidase with thrombospondin motifs 4 (ADAMTS4), a metallopeptidase expressed exclusively by OLs and dependent on adult OL genesis to maintain its expression in the ME. Both lack of Adamts4 and ADAMTS4 gain-of-function are associated with impaired glucose homeostasis and remodelling of the blood-hypothalamus barrier, indicating that optimal ADAMTS4 expression is required for the integrity of vascular permeability and normal glycaemic control. Finally, we show that ME ADAMTS4 expression is regulated by changes in peripheral glycaemia and is dysregulated in diabetes, providing a mechanism by which ME OLs contribute to the regulation of glucose homeostasis. | 2024-09-14 | |
Islet amyloid disrupts MHC Class II antigen presentation and protects NOD mice from autoimmune diabetes. | 10.1101/2024.09.10.612362 | Denroche, H. C.; Ng, V.; Velghe, J.; Suen, I.; Stanley, L.; Nackiewicz, D.; Komba, M.; Chen, S.; Soukhatcheva, G.; Dai, L.; Verchere, C. B. | Islet amyloid contributes to beta cell failure in type 2 diabetes through several mechanisms, one being the potent induction of local islet inflammation through activating inflammatory pathways in islet macrophages. We performed an unbiased phenotypic investigation of islet macrophages in the early stage of islet amyloid formation using single cell RNA sequencing of resident islet macrophages in mice with and without the amyloidogenic form of human islet amyloid polypeptide (hIAPP). This revealed that MHC Class II antigen presentation genes were strongly down-regulated in islet macrophages during islet amyloid formation. As islet amyloid has recently been reported in pancreases of people with type 1 diabetes, we sought to investigate the impact of islet amyloid in the NOD mouse model of type 1 diabetes. Both overexpression and physiological expression of hIAPP delayed diabetes in NOD mice relative to littermate controls, corresponding with decreased markers of antigen presentation and activation, as well as decreased immune cell infiltration in islets. Adoptive transfer studies showed that systemic autoimmune function remained intact and beta cells from hIAPP transgenic mice did not evade immune recognition by diabetogenic T cells, collectively indicating the protection from diabetes was mediated by localized disruption of antigen presentation in the pancreas. Consistent with this, incubation of dendritic cells with IAPP aggregates decreased MHC Class II surface expression and diminished antigen-specific T cell activation in vitro, through a phagocytosis-dependent mechanism. Collectively our data show that despite the well-established pro-inflammatory response of macrophages to IAPP aggregates, the uptake of IAPP aggregates during early amyloid formation also disrupts MHC Class II antigen presentation and slows beta cell autoimmunity. | 2024-09-14 | |
Single-cell and spatiotemporal transcriptomic profiling of brain immune infiltration following Venezuelan equine encephalitis virus infection | 10.1101/2024.09.12.612602 | Rangel, M.; Sebastian, A.; Leon, N.; Phillips, A.; Gorman, B.; Hum, N.; Weilhammer, D. R. | Neurotropic alphaviruses such as Venezuelan equine encephalitis virus (VEEV) are critical human pathogens that continually expand to naive populations and for which there are no licensed vaccines or therapeutics. VEEV is highly infectious via the aerosol route and is a recognized weaponizable biothreat that causes neurological disease in humans. The neuropathology of VEEV has been attributed to an inflammatory immune response in the brain yet the underlying mechanisms and specific immune cell populations involved are not fully elucidated. This study uses single-cell RNA sequencing to produce a comprehensive transcriptional profile of immune cells isolated from the brain over a time course of infection in a mouse model of VEEV. Analyses reveal differentially activated subpopulations of microglia, including a distinct type I interferon-expressing subpopulation. This is followed by the sequential infiltration of myeloid cells and cytotoxic lymphocytes, also comprising subpopulations with unique transcriptional signatures. We identify a subpopulation of myeloid cells that form a distinct localization pattern in the hippocampal region whereas lymphocytes are widely distributed, indicating differential modes of recruitment, including that to specific regions of the brain. Altogether, this study provides a high-resolution analysis of the immune response to VEEV in the brain and highlights potential avenues of investigation for therapeutics that target neuroinflammation in the brain. | 2024-09-14 | |
SARS-CoV-2 protein ORF3a induces Atg8ylation of lysosomal membranes | 10.1101/2024.09.12.612614 | Ajnar, D.; Sarkar, A.; Riyaz, S.; Menon, P.; Dutta, D.; Asati, P.; Sharma, P.; Pydi, S. P.; Kumar, S. | Autophagy Conjugation machinery forms a center piece of autophagy and is essential for sequestration of a broad range of cargo destined for degradation. Apart from its role in canonical autophagy, recent evidence suggests an unconventional role of conjugation machinery. Membrane Atg8ylation is one of the manifestations of autophagy, wherein ATG8 conjugation machinery recruit mammalian ATG8s (mATG8s) to the damaged membranes for repair or removal. Herein, we show that SARS-CoV-2 factor ORF3a induces membrane Atg8ylation and selectively inflicts lysophagy, a cellular response to evade apoptotic cell death. mATG8s and SNARE protein syntaxin 17 (STX17) interact with ORF3a and are required for Atg8ylation induced by ORF3a. ORF3a displaces mTOR from the lysosomes and affects nuclear translocation of TFEB, which is dependent on mATG8s and STX17. Despite mTOR inhibition, its conventional target ULK1 is dispensable for ORF3a induced Atg8ylation. In addition, mATG8s and STX17 protected against the cell death induced by ORF3a. Overall, our findings demonstrate ORF3a induced lysosomal membrane Atg8ylation while identifying the unexpected role of STX17 in Atg8ylation. | 2024-09-14 | |
Using correlative and mechanistic species distribution models to predict vector-borne disease risk for the current and future environmental and climatic change: a case study of West Nile Virus in the UK. | 10.1101/2024.09.12.612656 | Withers, A. J.; Croft, S.; Budgey, R.; Warren, D.; Johnson, N. | Globally, vector-borne diseases have significant impacts on both animal and human health, and these are predicted to increase with the effects of climate change. Understanding the drivers of such diseases can help inform surveillance and control measures to minimise risks both now and in the future. In this study, we illustrate a generalised approach for assessing disease risk combining species distribution models of vector and wildlife hosts with data on livestock and human populations using the potential emergence of West Nile Virus (WNV) in the UK as a case study. Currently absent in the UK, WNV is an orthoflavivirus with a natural transmission cycle between Culex mosquitos (Cx. pipiens and Cx. modestus) and birds. It can spread into non-target hosts (e.g., equids, humans) via mosquito bites where it can cause febrile disease with encephalitis and mortality in severe cases. We compared six correlative species distribution models and selected the most appropriate for each vector based on a selection of performance measures and compared this to mechanistic species distribution models and known distributions. We then combined these with correlative species distribution models of representative avian hosts, equines, and human population data to predict risk of WNV occurrence. Our findings highlighted areas at greater risk of WNV due to higher habitat suitability for both avian hosts and vectors, and considered how this risk could change by 2100 under a best-case Shared Socioeconomic Pathway (SSP1) and worst-case (SSP5) future climate scenario. Generally, WNV risk in the future was found to increase in south-eastern UK and decrease further north. Overall, this paper presents how current and future vector distributions can be modelled and combined with projected host distributions to predict areas at greater risk of novel diseases. This is important for policy decision making and contingency preparedness to enable adaptation to changing environments and the resulting shifts in vector-borne diseases that are predicted to occur. | 2024-09-14 | |
Assessing the risk of highly pathogenic avian influenza H5 viruses spread within French broiler and layer production networks | 10.1101/2024.09.11.612235 | Hautefeuille, C.; Munoz, F.; Dauphin, G.; Paul, M.; Peyre, M.; Goutard, F. | Since 2015, French poultry production is threatened almost every year by a reintroduction of highly pathogenic avian influenza H5 viruses. The duck sector was the most concerned by this crisis but other sectors such as broiler, layer and turkey were also affected by outbreaks. The objective of this work was to assess the risk of highly pathogenic avian influenza H5 virus transmission from one farm to another within the French broiler and layer production network. This study used the WOAH risk assessment framework. After drawing up a scenario tree of virus transmission from one farm to another, data were collected through a literature review or through experts' elicitation. Three questionnaires were developed according to the experts' field of expertise: avian influenza, broiler and layer sectors. The experts' estimates were combined using a beta distribution weighted by their confidence level. A Monte Carlo iteration process was used to combine the different probabilities of the scenario tree and to assess the transmission risk. In the broiler sector, the highest transmission probabilities were observed if the exposed farm was an indoor broiler farm and the source a broiler farm (indoor or free-range). The high transmission probability between broiler farm integrated within the same company highlighted the role of integration in this probability. In the layer sector, the highest transmission probabilities were observed if the source farm was a free-range farm and the exposed farm a production farm (indoor or free-range). The sensitivity analysis showed that the exposed farm's biosecurity had a significant impact on the transmission probability. Our results provide an insight on the role of each type of farms in the virus spread within the French broiler and layer production network and will be useful for the implementation of control measures such as movement restriction or vaccination. | 2024-09-14 | |
The Draparnaldia genome: alternative mechanisms for multicellularity and terrestrialization in green plants | 10.1101/2024.09.12.612648 | Caisova, L.; Crombez, E.; Arellano, M. S. T.; Gut, M.; Alioto, T. S.; Gomez-Garrido, J.; Dabad, M.; Esteve-Codina, A.; Petrik, I.; Pencik, A.; Novak, O.; Van de Peer, Y.; Vicoso, B.; Friml, J. | Green plants contain two algal lineages: Streptophyte algae that diverged into land plants and Chlorophyte algae that are mostly aquatic. Draparnaldia is a Chlorophyte alga morphologically resembling mosses and living in both, the aquatic and terrestrial habitats. Because of its complex morphology and terrestrial adaptations, Draparnaldia can provide new insights into the evolution of multicellularity and terrestrialization in green plants. To develop Draparnaldia into a model, we de novo sequenced its genome and transcriptomes, and profiled its phytohormone repertoire. We found that 1) Expanded gene families in Draparnaldia with respect to unicellular Chlamydomonas are linked to multicellularity and abiotic stresses. 2) Draparnaldias terrestrial adaptations are reflected at both the morphological and molecular levels. 3) Draparnaldia synthesizes most of the phytohormones used by land plants to thrive in terrestrial habitats. All of this makes Draparnaldia a powerful model to uncover and study alternative evolutionary trajectories towards multicellularity and terrestrialization in plants. | 2024-09-14 | |
EBAX-1/ZSWIM8 destabilizes miRNAs resulting in transgenerational memory of a predatory trait | 10.1101/2024.09.10.612280 | Quiobe, S. P.; Kalirad, A.; Roeseler, W.; Witte, H.; Wang, Y.; Roedelsperger, C.; Sommer, R. J. | Environmental influences on traits and associated transgenerational epigenetic inheritance have widespread implications, but remain controversial and underlying mechanisms poorly understood. We introduce long-term environmental induction experiments on alternative diets in Pristionchus pacificus, a nematode exhibiting mouth-form plasticity including predation, by propagating 110 isogenic lines for 101 generations with associated food-reversal experiments. We found dietary induction and subsequent transgenerational memory of the predatory morph and identified a role of ubiquitin ligase EBAX-1/ZSWIM8 in this process. Ppa-ebax-1 mutants have no memory and Ppa-EBAX-1 destabilizes the clustered microRNA family miR-2235a/miR-35. Deletions of a cluster of 44 identical miR-2235a copies resulted in precocious and extended transgenerational inheritance of the predatory morph. These findings indicate that EBAX-1/ZSWIM8 destabilizes miRNAs resulting in transgenerational memory, suggesting a role for target-directed miRNA degradation. | 2024-09-14 | |
LncRNA TAAL is a Modulator of Tie1-Mediated Vascular Function in Diabetic Retinopathy | 10.1101/2024.09.13.612383 | Ranjan, G.; Arora, S.; Sharma, S.; Sharma, L.; Bhoyar, R. C.; Senthivel, V.; Scaria, V.; Chakrabarti, S.; Kaur, I.; Motiani, R. K.; Sivasubbu, S. | Diabetic retinopathy (DR) is a leading cause of vision impairment and blindness resulting from abnormal retinal vascular changes due to chronic hyperglycemia. The ANG-TIE signaling pathway, crucial for vascular growth and remodeling, is a key therapeutic target in DR. Through a protein-centric approach, we identified a novel lncRNA an named it Tie1-associated angiogenic lncRNA (TAAL), which regulates endothelial cell migration, proliferation, tube formation, permeability, and angiogenesis via changing calcium and cytoskeleton dynamics in endothelial cells. In zebrafish, modulation of the TAAL orthologue led to angiogenic defects, which were rescued by human TAAL. Molecular studies revealed that TAAL negatively regulates Tie1 protein expression through ubiquitin-mediated degradation. Notably, TAAL is upregulated in the blood of DR patients and is reduced in endothelial DR cell models. Overexpression of TAAL restores permeability and VE-cadherin surface expression. These findings present a novel function of lncRNA TAAL as a critical regulator of Tie1 protein turnover with potential therapeutic applications for diabetic retinopathy. | 2024-09-14 | |
Haplotype-resolved assemblies provide insights into genomic makeup of the oldest grapevine cultivar (Munage) in Xingjiang | 10.1101/2024.09.11.612401 | Zhong, H.; Shi, x.; Zhang, F.; Wang, X.; yadav, v.; Zhou, X.; Cao, s.; Zhang, S.; Zhang, C.; Qiao, J.; Liu, Z.; Zhang, y.; Liu, y.; Wang, H.; Xue, h.; Zhang, M.; Zhang, T.-H.; Zhou, Y.; Wu, X.; Xiao, H. | Munage, an ancient grape variety that has been cultivated for thousands of years in Xinjiang, China, is recognized for its exceptional fruit traits. There are two main types of Munage: white fruit (WM) and red fruit (RM). However, the lack of a high-quality genomic resources has impeded effective breeding and restricted the potential for expanding these varieties to other growing regions. In this study, we assembled haplotype-resolved genome assemblies for WM and RM, alongside integrated whole genome resequencing (WGS) data and transcriptome data to illuminate specific mutations and associated genes in Munake and the genes associated with fruit color traits. Selective analysis between Munage clones and Eurasian grapes suggested that adaptive selection exists in Munage grapes, with genes enriched in processes including cell maturation, plant epidermal cell differentiation, and root epidermal cell differentiation. The study examined the mutations within Munage grapes and found that the genes PMAT2 on chromosome 12 and MYB123 on chromosome 13 are likely responsible for color variation in RM. These findings provide crucial genetic resources for investigating the genetics of the ancient Chinese grape variety, Munage, and will facilitate the genetic improvement in grapevine. | 2024-09-14 | |
DiatOmicBase, a gene-centered platform to mine functional omics data across diatom genomes | 10.1101/2024.09.12.612655 | Villar, E.; Zweig, N.; Vincens, P.; Cruz de Carvalho, H.; Duchene, C.; Liu, S.; Monteil, R.; Dorrell, R. G.; Fabris, M.; Vandepoele, K.; Bowler, C.; Falciatore, A. | Diatoms are prominent microalgae found in all aquatic environments. Over the last 20 years, thanks to the availability of genomic and genetic resources, diatom species such as Phaeodactylum tricornutum have emerged as valuable experimental model systems for exploring topics ranging from evolution to cell biology, (eco)physiology and biotechnology. Since the first genome sequencing in 2008, numerous genome-enabled datasets have been generated, based on RNA-Seq and proteomics, epigenomes, and ecotype variant analysis. Unfortunately, these resources, generated by various laboratories, are often in disparate formats and challenging to access and analyze. Here we present DiatOmicBase, a genome portal gathering comprehensive omics resources from P. tricornutum and two other diatoms to facilitate the exploration of dispersed public datasets and the design of new experiments based on the prior-art. DiatOmicBase provides gene annotations, transcriptomic profiles and a genome browser with ecotype variants, histone and methylation marks, transposable elements, non-coding RNAs, and read densities from RNA-Seq experiments. We developed a semi-automatically updated transcriptomic module to explore both publicly available RNA-Seq experiments and users private datasets. Using gene-level expression data, users can perform exploratory data analysis, differential expression, pathway analysis, biclustering, and co-expression network analysis. Users can create heatmaps to visualize precomputed comparisons for selected gene subsets. Automatic access to other bioinformatic resources and tools for diatom comparative and functional genomics is also provided. Focusing on the resources currently centralized for P. tricornutum, we showcase several examples of how DiatOmicBase strengthens molecular research on diatoms, making these organisms accessible to a broad research community. | 2024-09-14 | |
The level of endothelial glycocalyx maturity modulates interactions with charged nano-materials | 10.1101/2024.09.10.611831 | Bridges, C.; Fu, L.; Yeow, J.; Huang, X.; Jackson, M.; Kuchel, R.; Sterling, J.; Baker, S.; Lord, M. | Nanomaterials have been extensively investigated for their potential in delivering therapeutics to target tissues, but few have advanced to clinical application. The luminal surface of endothelial cells that line blood vessels are covered by a glycocalyx, a complex extracellular matrix rich in anionic glycans. However, the role of this glycocalyx in governing nanomaterial- cell interactions is often overlooked. In this study, we demonstrate that gold nanoparticles functionalized with branched polyethyleneimine (AuNP+) bind to primary human endothelial cells expressing either a developing or mature glycocalyx, with the interaction involving hyaluronan and heparan sulfate. Notably, the mature glycocalyx decreases the toxicity of AuNP+. In contrast, lipoic acid-functionalized gold nanoparticles (AuNP-) bind to endothelial cells with a developing glycocalyx, but not a mature glycocalyx. To further investigate this phenomenon, we studied charged polymers, including poly(arginine) (polyR) and poly(glutamic acid) (polyE). PolyE does not associate with endothelial cells regardless of glycocalyx maturity, but when glycans are enzymatically degraded, it can bind to the cells. Conversely, polyR associates with endothelial cells irrespective of glycocalyx maturity or glycan degradation. These findings highlight the intricate relationship between nanomaterial charge and presentation in interactions with endothelial cells, offering insights for modulating nanomaterial interactions with the blood vessel wall. | 2024-09-14 | |
Causality Analysis of Protein Corona Composition: Phosphatidylcholine-Enhances Plasma Proteome Profiling by Proteomics | 10.1101/2024.09.10.612356 | Rafieioskouei, A.; Rogale, K.; Dibavar, A. S.; Mahmoudi, M.; Bonakdarpour, B. | The study of the protein corona, the immediate and evolving biomolecular coating that forms on the surface of nanoparticles when exposed to a biological environment, is a crucial area in nanomedicine. This phenomenon significantly influences the behavior, functionality, and biological interactions of nanoparticles with biosystems. Until now, conclusions regarding the role of the protein corona in specific biological applications have been based on establishing correlation rather than causation. By understanding causality, researchers can predict how changes in nanoparticle properties or biological conditions will affect protein corona composition, in turn affecting the nanoparticle interactions with the biosystems and their applications. This predictive capability is essential for designing nanoparticles with specific characteristics tailored for therapeutic and diagnostic nanomedicine applications. Here, we explore the concept of actual causality (by Halpern and Pearl) to mathematically prove how various small molecules, including metabolites, lipids, vitamins, and nutrients, spiked into plasma can induce diverse protein corona patterns on identical nanoparticles. This approach significantly enhances the depth of plasma proteome profiling. Our findings reveal that among the various spiked small molecules, phosphatidylcholine was the actual cause of the observed increase in the proteomic depth of the plasma sample. By considering the concept of causality in the field of protein corona, the nanomedicine community can substantially improve their ability to design safer and more efficient nanoparticles for both diagnostic and therapeutic purposes. | 2024-09-14 | |
ScReNI: single-cell regulatory network inference through integrating scRNA-seq and scATAC-seq data | 10.1101/2024.09.10.612385 | Xu, X.; Liang, Y.; Tang, M.; Wang, J.; Wang, X.; Li, Y.; Wang, J. | Single cells exhibit heterogeneous gene expression profiles and chromatin accessibility, measurable separately via single-cell RNA sequencing (scRNA-seq) and single-cell transposase chromatin accessibility sequencing (scATAC-seq). Consequently, each cell possesses a unique gene regulatory network. However, limited methods exist for inferring cell-specific regulatory networks, particularly through the integration of scRNA-seq and scATAC-seq data. Here, we develop a novel algorithm named single-cell regulatory network inference (ScReNI), which leverages k-nearest neighbors and random forest algorithms to integrate scRNA-seq and scATAC-seq data for inferring gene regulatory networks at the single-cell level. ScReNI is built to analyze both paired and unpaired datasets for scRNA-seq and scATAC-seq. Using these two types of single-cell sequencing datasets, we validate that a higher fraction of regulatory relationships inferred by ScReNI are detected by chromatin immunoprecipitation sequencing (ChIP-seq) data. ScReNI shows superior performance in network-based cell clustering when compared to existing single-cell network inference methods. Importantly, ScReNI offers the unique function of identifying cell-enriched regulators based on each cell-specific network. In summary, ScReNI facilitates the inferences of cell-specific regulatory networks and cell-enriched regulators. | 2024-09-14 | |
LoVis4u: Locus Visualisation tool for comparative genomics | 10.1101/2024.09.11.612399 | Egorov, A. A.; Atkinson, G. C. | Summary: Comparative genomic analysis often involves visualisation of alignments of genomic loci. While several software tools are available for this task, ranging from Python and R libraries to standalone graphical user interfaces, there is lack of a tool that offers fast, automated usage and the production of publication-ready vector images. Here we present LoVis4u, a command-line tool and Python API designed for highly customizable and fast visualisation of multiple genomic loci. LoVis4u generates vector images in PDF format based on annotation data from GenBank or GFF files. It is capable of visualising entire genomes of bacteriophages as well as plasmids and user-defined regions of longer prokaryotic genomes. Additionally, LoVis4u offers optional data processing steps to identify and highlight accessory and core genes in input sequences. Availability and Implementation: LoVis4u is implemented in Python3 and runs on Linux and MacOS. The command-line interface covers most practical use cases, while the provided Python API allows usage within a Python program, integration into external tools, and additional customisation. Source code is available at the GitHub page: github.com/art-egorov/lovis4u. Detailed documentation that includes an example-driven guide is available from the software home page: art-egorov.github.io/lovis4u. | 2024-09-14 | |
SVbyEye: A visual tool to characterize structural variation among whole genome assemblies | 10.1101/2024.09.11.612418 | Porubsky, D.; Guitart, X.; Yoo, D.; Dishuck, P. C.; Harvey, W. T.; Eichler, E. E. | Motivation We are now in the era of being able to routinely generate highly contiguous (near telomere-to-telomere) genome assemblies of human and nonhuman species. Complex structural variation and regions of rapid evolutionary turnover are being discovered for the first time. Thus, efficient and informative visualization tools are needed to evaluate and directly observe structural differences between two or more genomes. Results We developed SVbyEye, an open-source R package to visualize and annotate sequence-to-sequence alignments along with various functionalities to process alignments in PAF format. The tool facilitates the characterization of complex SVs in the context of sequence homology helping resolve the mechanisms underlying their formation. Availability and implementation SVbyEye is available at https://github.com/daewoooo/SVbyEye. | 2024-09-14 | |
A high-throughput phenotypic screen combined with an ultra-large-scale deep learning-based virtual screening reveals novel scaffolds of antibacterial compounds | 10.1101/2024.09.11.612340 | Scalia, G.; Rutherford, S. T.; Lu, Z.; Buchholz, K. R.; Skelton, N.; Chuang, K.; Diamant, N.; Huetter, J.-C.; Luescher, J.; Miu, A.; Blaney, J.; Gendelev, L.; Skippington, E.; Zynda, G.; Dickson, N.; Koziarski, M.; Bengio, Y.; Regev, A.; Tan, M.-W.; Biancalani, T. | The proliferation of multi-drug-resistant bacteria underscores an urgent need for novel antibiotics. Traditional discovery methods face challenges due to limited chemical diversity, high costs, and difficulties in identifying structurally novel compounds. Here, we explore the integration of small molecule high-throughput screening with a deep learning-based virtual screening approach to uncover new antibacterial compounds. Leveraging a diverse library of nearly 2 million small molecules, we conducted comprehensive phenotypic screening against a sensitized Escherichia coli strain that, at a low hit rate, yielded thousands of hits. We trained a deep learning model, GNEprop, to predict antibacterial activity, ensuring robustness through out-of-distribution generalization techniques. Virtual screening of over 1.4 billion compounds identified potential candidates, of which 82 exhibited antibacterial activity, illustrating a 90X improved hit rate over the high-throughput screening experiment GNEprop was trained on. Importantly, a significant portion of these newly identified compounds exhibited high dissimilarity to known antibiotics, indicating promising avenues for further exploration in antibiotic discovery. | 2024-09-14 | |
The Survival of β-lactoglobulin Peptides in the Archaeological Record: Vulnerability vs. Sequence Variation | 10.1101/2024.09.13.612646 | Fonseca, B.; Freeman, C. L.; Collins, M. J. | It is a strange observation, given the cultural co-evolution of dairying, that milk proteins are more commonly reported than any other food proteins in the archaeological record. The whey protein {beta}-lactoglobulin and in particular its eleven amino acid long peptide T125PEVDXEALEK135 seems to be preferentially preserved in both ceramic vessels and teeth (dental calculus). An amino acid substitution in the middle of the chain is valuable to track livestock management because it permits differentiation between key animal species used in dairying. The persistence of this peptide is, however, unusual as its acidic nature makes it more vulnerable to hydrolysis. Moreover, selection for the ability to digest raw milk - more specifically, the continued production of the milk sugar enzyme lactase beyond the age of normal weaning - did not begin until the early Bronze Age. It is therefore unclear why it is a milk peptide, in particular a peptide associated with the lactose-rich whey fraction, that is one of the most commonly recovered dietary peptides. The unexpected preservation of T125PEVDXEALEK135 thus presents a good case study to uncover patterns of protein survival in the archaeological record. We have previously explored the dynamics of the bovine variation of the peptide (X=Asp130) and its likelihood to undergo hydrolysis in solution. In this study, we turn our attention to the ovine (X=Asn130) and the caprine (X=Lys130) variations of the {beta}-lactoglobulin peptide to determine how the mutation in the amino acid in position 6 affects peptide conformations and vulnerability in bulk water. To do this, we use Molecular Dynamics as implemented in GROMACS 2020, with the Amber14SB forcefield and the SPC/E water model. We first perform extensive conformational analysis of both peptides in solution to determine stable structures. Then, using analogous methodology to that developed in our earlier study of the bovine peptide, we identify geometric arrangements between water and peptide that may be more prone to hydrolysis. | 2024-09-14 | |
Possible Mechanisms Of Action Of Light Inert Gases On Chemiluminescence Arising As A Result Of Lipid Peroxidation | 10.1101/2024.09.12.612290 | Oliynyk, I. | The use of inert gases in biology and medicine and their effect on biological objects both in vitro and in vivo remains an active area of research. It has been established that light noble gases affect antioxidant processes, free radical oxidation, and enhance chemiluminescence, but an explanation of the physical and chemical mechanisms of this effect is still lacking and is key to further theoretical and experimental studies, given the broad prospects for the use of noble gases in medicine. In this article, we present two of the possible mechanisms of light inert gases' effect on chemiluminescence (CL), a phenomenon that occurs as a result of free radical recombination and chain breakage during lipid peroxidation. Since the effect on oxidation, in turn, precedes the effect on the antioxidant system and the body's defense mechanisms. One of the mechanisms of influence is based on the ability of inert gases to dissolve well in lipids and dissolve poorly in water. Their ability to dissolve in lipid bilayers and affect the conformation of lipid complexes can increase the surface area available for oxidation, the surface area that absorbs radiation and reduce the density of the environment, potentially increasing the availability of oxygen for oxidation reactions. This is the so-called spatial mechanism of inert gas influence on oxidation and chemiluminescence. The second mechanism is based on the influence on the quantum chemical parameters of the reaction medium. The acceleration of VT relaxation processes, the impact on the components of the medium in quenching excited states, and the radiative decay time of the excited state. | 2024-09-14 | |
BST2 induces vascular smooth muscle cell plasticity and phenotype switching during cancer progression | 10.1101/2024.09.10.612298 | Bell, C.; Baylis, R.; Lopez, N.; Ma, W. F.; Gao, H.; Wang, F.; Bamezai, S.; Fu, C.; Kojima, Y.; Adkar, S.; Luo, L.; Miller, C.; Leeper, N. L. | Background: Smooth muscle cell (SMC) plasticity and phenotypic switching play prominent roles in the pathogenesis of multiple diseases, but their role in tumorigenesis is unknown. We investigated whether and how SMC diversity and plasticity plays a role in tumor angiogenesis and the tumor microenvironment. Methods and Results: We use SMC-specific lineage-tracing mouse models and single cell RNA sequencing to observe the phenotypic diversity of SMCs participating in tumor vascularization. We find that a significant proportion of SMCs adopt a phenotype traditionally associated with macrophage-like cells. These cells are transcriptionally similar to resolution phase M2b macrophages, which have been described to have a role in inflammation resolution. Computationally predicted by the ligand-receptor algorithm CellChat, signaling from BST2 on the surface of tumor cells to PIRA2 on SMCs promote this phenotypic transition; in vitro SMC assays demonstrate upregulation of macrophage transcriptional programs, and increased proliferation, migration, and phagocytic ability when exposed to BST2. Knockdown of BST2 in the tumor significantly decreases the transition towards a macrophage-like phenotype, and cells that do transition have a comparatively higher inflammatory signal typically associated with anti-tumor effect. Conclusion: As BST2 is known to be a poor prognostic marker in multiple cancers where it is associated with an M2 macrophage-skewed TME, these studies suggest that phenotypically switched SMCs may have a previously unidentified role in this immunosuppressive milieu. Further translational work is needed to understand how this phenotypic switch could influence the response to anti-cancer agents and if targeted inhibition of SMC plasticity would be therapeutically beneficial. | 2024-09-14 | |
On the design and stability of cancer adaptive therapy cycles: deterministic and stochastic models | 10.1101/2024.09.10.612338 | Vilela, Y. G.; Fassoni, A. C.; Neves, A. G. M. | Adaptive therapy is a promising paradigm for treating cancers, that exploits competitive interactions between drug-sensitive and drug-resistant cells, thereby avoiding or delaying treatment failure due to evolution of drug resistance within the tumor. Previous studies have shown the mathematical possibility of building cyclic schemes of drug administration which restore tumor composition to its exact initial value in deterministic models. However, algorithms for cycle design, the conditions on which such algorithms are certain to work, as well as conditions for cycle stability remain elusive. Here, we state biologically motivated hypotheses that guarantee existence of such cycles in two deterministic classes of mathematical models already considered in the literature: Lotka-Volterra and adjusted replicator dynamics. We stress that not only existence of cyclic schemes, but also stability of such cycles is a relevant feature for applications in real clinical scenarios. We also analyze stochastic versions of the above deterministic models, a necessary step if we want to take into account that real tumors are composed by a finite population of cells subject to randomness, a relevant feature in the context of low tumor burden. We argue that the stability of the deterministic cycles is also relevant for the stochastic version of the models. In fact, Dua, Ma and Newton [Cancers (2021)] and Park and Newton [Phys. Rev. E (2023)] observed breakdown of deterministic cycles in a stochastic model (Moran process) for a tumor. Our findings indicate that the breakdown phenomenon is not due to stochasticity itself, but to the deterministic instability inherent in the cycles of the referenced papers. We then illustrate how stable deterministic cycles avoid for very large times the breakdown of cyclic treatments in stochastic tumor models. | 2024-09-14 | |
Metastatic tumor cells in bone marrow differ from paired neuroblastoma tumor and contain subsets with therapy-resistant characteristics | 10.1101/2024.09.13.612231 | Hochheuser, C.; Boltjes, A.; Keller, K. M.; Tol, S.; van de Mheen, M.; Pita Barros, C.; van Gestel-Fadaie, Z.; van Kuilenburg, A. B. P.; van Hooff, S.; Voermans, C.; Molenaar, J. J.; Tytgat, G. A. M.; Timmerman, I. | Bone marrow (BM) is a common site for solid tumor metastasis, often causing poor outcome. Here, we define the characteristics of BM-disseminated tumor cells (DTCs) using neuroblastoma as a model. We combined single-cell RNA-sequencing (scRNA-seq) and cell-surface protein analysis using 7 paired BM and primary tumor (PT) samples and found that DTCs contain a higher percentage of cycling cells and higher expression of neurodevelopmental genes compared to corresponding PT cells. In 6 patients, the copy number variation profile differed between PT cells and DTCs, indicating spatial heterogeneity. Within the BM, we detected dormant DTCs with potentially reduced chemosensitivity; this population contained cells expressing low levels of the immunotherapeutic antigen GD2 and increased NGFR expression. In conclusion, we characterized DTCs that are particularly challenging to target, offering new avenues for developing therapeutic strategies designed to target all subpopulations within the highly complex metastatic site, thereby preventing the development of drug-resistant clones. | 2024-09-14 | |
Predicting Task Activation Maps from Resting-State Functional Connectivity using Deep Learning | 10.1101/2024.09.10.612309 | Madsen, S. J.; Uddin, L. Q.; Mumford, J. A.; Barch, D. M.; Fair, D. A.; Gotlib, I. H.; Poldrack, R. A.; Kuceyeski, A.; Saggar, M. | Recent work has shown that deep learning is a powerful tool for predicting brain activation patterns evoked through various tasks using resting state features. We replicate and improve upon this recent work to introduce two models, BrainSERF and BrainSurfGCN, that perform at least as well as the state-of-the-art while greatly reducing memory and computational footprints. Our performance analysis observed that low predictability was associated with a possible lack of task engagement derived from behavioral performance. Furthermore, a deficiency in model performance was also observed for closely matched task contrasts, likely due to high individual variability confirmed by low test-retest reliability. Overall, we successfully replicate recently developed deep learning architecture and provide scalable models for further research. | 2024-09-14 | |
Boosting forward connectivity between primary visual and body selective cortex reduces interference of task irrelevant dimensions on body judgements | 10.1101/2024.09.10.612296 | Gandolfo, M.; D'argenio, G.; Downing, P. E.; Urgesi, C. | We effortlessly categorise other people along socially relevant categories such as sex, age, and emotion. A core question in social vision relates to whether we perceive these categories independently or in relation to each other. Here, we investigated categorisation of sex and emotion from the body, finding that participants generally fail to fully ignore task-irrelevant variations of sex while judging body emotional expressions. In contrast, sex categorisation was unaffected by variations in emotional expression. We propose that this asymmetric interaction between sex and emotion may arise because of bottom-up visual processing, due to partially shared visual features used for both judgments, or because of top-down, categorical associations between sex and emotion categories. To disentangle these possibilities, we used cortico-cortical paired associative stimulation (ccPAS) to modulate the connectivity between primary visual cortex and the extrastriate body area. We posited that boosting forward connectivity between these regions would increase efficiency of feature-based processing, while boosting feedback connectivity would enhance the separability of semantic categories related to sex and emotion. We found that boosting forward connectivity eliminated the interference of the irrelevant variation of body sex on body emotion judgments, while the same interference remained unaffected with modulation of feedback connectivity. These findings suggest that interactions between sex and emotion in body perception emerge during the perceptual analysis of the stimuli, and add to our understanding of person perception and social categorization. | 2024-09-14 | |
ARHGEF3 Regulates Hair Follicle Morphogenesis | 10.1101/2024.09.13.612256 | Laurin, M.; Kalyanakrishnan, K.; Beaudin, A.; Jette, A.; Ghezelbash, S.; Ioana Hotea, D.; Chen, J.; Lefrancois, P. | During embryogenesis, cells arrange into precise patterns that enable tissues and organs to develop specialized functions. Despite its critical importance, the molecular choreography behind these collective cellular behaviors remains elusive, posing a major challenge in developmental biology and limiting advances in regenerative medicine. By using the mouse hair follicle as a mini-organ system to study the formation of bud-like structures during embryonic development, our work uncovers a crucial role for the Rho GTPase regulator ARHGEF3 in hair follicle morphogenesis. We demonstrate that Arhgef3 expression is upregulated at the onset of hair follicle placode formation. In Arhgef3 knockout animals, we observed defects in placode cell compaction, leading to impaired hair follicle downgrowth. Through cell culture models, we show that ARHGEF3 promotes F-actin accumulation at the cell cortex and P-cadherin enrichment at cell-cell junctions. Collectively, our study identifies ARHGEF3 as a new regulator of cell shape rearrangements during hair placode morphogenesis, warranting further exploration of its role in other epithelial appendages that arise from similar developmental processes. | 2024-09-14 | |
Beneficial rhizobacteria and virus infection modulate the soybean metabolome and influence the feeding preferences of the virus vector Epilachna varivestis. | 10.1101/2024.09.11.612434 | Pulido, H.; De Moraes, C. M.; Mauck, K.; Mescher, M. C. | There is growing evidence that microbial plant symbionts shape interactions between plants and other organisms by modulating gene expression and metabolism. However, the detailed mechanisms mediating such effects are not well understood, particularly in systems where plants interact simultaneously with multiple symbionts and antagonists. In this study, we employed a multi-factorial design to explore the individual and combined effects of two plant-beneficial rhizobacteria (Delftia acidovorans and Bradyrhizobium japonicum) and a pathogen (Bean pod mottle virus: BPMV) on gene expression and metabolite production by soybean plants, as well as downstream effects on plant interactions with a beetle vector of BPMV Epilachna varivestis. Our results document microbial effects on basic metabolism and defense pathways, resulting in increased levels of primary metabolites and depletion of secondary metabolites. These changes are consistent with the observed feeding preferences of beetles for rhizobia-inoculated and virus-infected plants. Together, our results indicate that BPMV infection and rhizobacteria colonization cause dramatic changes in plant metabolites related to nutrition and defense, with significant consequences for an agriculturally important pathosystem. | 2024-09-14 | |
Foot placement control underlies stable locomotion across species | 10.1101/2024.09.10.612345 | De Comite, A.; Seethapathi, N. | Animals navigate their environment stably without inefficient course corrections despite unavoidable errors. In humans and some robots, this stability is achieved by controlling the placement of the foot on the ground such that recent movement errors are corrected. However, it is unknown whether and how animals with diverse nervous systems and body mechanics use such foot placement control: foot trajectories of many-legged animals are considered as stereotypical velocity-driven patterns, as opposed to error-driven. Here, we posit a control structure for stabilizing foot placement in any legged embodiment by unifying velocity-driven and body state-driven contributions, and develop a framework to discover control strategies used across species from natural locomotor variability. Using this framework, we find evidence for body state-dependent foot placement control in flies and mice, previously only shown to exist in humans. We discover that the urgency and centralization of the foot placement control strategy is shaped by the animal's neuromechanical embodiment. More inherently stable many-legged embodiment is associated with a lower control magnitude and timescale. Further, many-legged embodiment is accompanied by decentralized control with modular control functions, timescales, and gains, whereas analogous functions are centralized across both legs in humans. Our approach discovers signatures of stabilizing control across species and reveals how different neuromechanical embodiments achieve a shared functional goal: foot placement control. | 2024-09-14 | |
Evaluation of stress tolerance in IR64 rice near-isogenic lines carrying SUB1 and DRO1 genes | 10.1101/2024.09.11.612414 | Sakagami, J.-I.; Soe, I.; Odama, E.; Chepkoech, R.; Ookawa, T.; Ismail, A. M. | Flooding and drought significantly reduce rice yield in rainfed environments. Rice varieties that tolerate complete inundation for up to two weeks carry the SUB1A gene, while those enduring deeper water conditions for weeks or months have the SK1 and SK2 genes. Conversely, the DRO1 gene, responsible for deep-rooting, helps in water acquisition under drought. In this study, we compared the growth of NIL-SUB1DRO1 a novel rice genotype with an IR64 background regarding its dual tolerance to submergence and drought. Additionally, we assessed its recovery capacity after exposure to stress. Sixteen and thirteen-days old seedlings of three genotypes (Experiment 1-1 and 2-1) and fourteen-days old seedlings of the two genotypes (Experiment 1-2 and 2-2) were tested under submergence and drought stress in a greenhouse experimental condition respectively. Seedlings were submerged for 10 and 7 days and then allowed to recover for 10 and 7 days respectively. In the drought experiment, seedlings underwent 29 days of drought (Experiment 2-1) and 18 days of drought, followed by 10 days of rewatering (Experiment 2-2). Growth parameters were measured before and after treatment, 4 days after submergence in experiment 1-1 and after the recovery periods. Submergence and drought adversely affected growth and performance. Shoot elongation in submerged plants was reduced by 29.2% for NIL-SUB1DRO1 compared to IR64. Accelerated shoot elongation of IR64 negatively affected its recovery. Chlorophyll content and maximum fluorescence of IR64 were significantly lower than other genotypes after 10 and 7 days of complete submergence. Ten days after recovery in experiment 1-1 the Chlorophyll content and maximum fluorescence of IR64 were not significantly different compared to other genotypes. Seven days after of recovery in experiment 1-2, NIL-SUB1DRO1 had significantly higher chlorophyll content and maximum fluorescence than IR64. After 29 days of drought the tiller number and leaf area of IR64 was lower than other genotypes (Experiment 2-1), while in Experiment 2-2 during drought treatment and recovery, NIL-SUB1DRO1 had greater relative water content, leaf water potential, leaf area, SPAD value, dry weights of shoots and roots, root length, surface area and volume compared to IR64. stomatal conductance of IR64 was higher than NIL-SUB1DRO1 during drought, leading to greater water loss and reduced growth during recovery. NIL-SUB1DRO1 absorbed and retained water more effectively under dry conditions. NIL-SUB1DRO1 and NIL-SUB1 is tolerant to submergence and NIL-SUB1DRO1 and NIL-DRO1 to drought, with no negative effects from combining these genes in modern rice varieties for rainfed lowlands. | 2024-09-14 | |
Nutritional analysis of commercially available, complete plant- and meat-based dry dog foods in the UK | 10.1101/2024.09.11.612409 | Brociek, R. A.; Li, D.; Broughton, R.; Gardner, D. S. | Background: Adoption of a plant-based diet is a popular lifestyle choice for many owners of canine companion animals. Increasingly, owners would like to feed their canine companions a similar diet. A plant-based dietary pattern has been reported to be associated with some micronutrient deficiencies. Complete dog foods are, by definition, supposed to be nutritionally replete in all macro- and micronutrients. Few studies have reported a full nutritional analysis of complete, dry plant- versus meat-based dog foods. Method: Here, 31 pet foods (n=19 meat-based, n=6 veterinary and n=6 plant-based) were analysed for total protein content and individual amino acids, fatty acids, major and trace elements, vitamin D and all B-vitamins. Results: Nutritional composition of meat and plant-based foods were similar, except for iodine and B-vitamins, which were lower in plant-based foods. The majority (66%) of veterinary diets with lower total protein by design, were also deficient in one or more essential amino acids. Isolated instances of non-compliance to nutritional guidelines were observed across all food-groups. Of the tested nutrients 55%, 16%, 24% and 100% of foods met all amino acid, mineral, B-vitamin, and vitamin D guidelines, respectively. Conclusions: Adopting a plant-based dietary pattern for your companion canine can provide nutritional adequacy with respect to the majority of macro- and micronutrients, but feeding supplemental iodine and B-vitamins should be considered. Veterinary diets, purposely low in crude protein, often have less than optimal essential amino acid composition. These data provide important new information for owners of companion canines being fed plant-based or veterinary diets. | 2024-09-14 | |
Pro-inflammatory mediators sensitise Transient Receptor Potential Melastatin 3 cation channel (TRPM3) signalling in mouse sensory neurons | 10.1101/2024.09.11.612393 | Aguilera-Lizarraga, J.; Lim, T. K.; Pattison, L. A.; Paine, L. W.; Bulmer, D. C.; Smith, E. S. J. | Pro-inflammatory mediators can directly activate pain-sensing neurons, known as nociceptors. Additionally, these mediators can potentiate or sensitise ion channels and receptors expressed by these cells through transcriptional and post-translational modulation, leading to nociceptor hypersensitivity. A well-characterised group of ion channels that subserve nociceptor sensitisation is the transient receptor potential (TRP) superfamily of cation channels. For example, the roles of TRP channels vanilloid 1 (TRPV1) and ankyrin 1 (TRPA1) in nociceptor sensitisation and inflammatory pain have been extensively documented. In the case of TRP melastatin 3 (TRPM3), however, despite the increasing recognition of this channel\'s role in inflammatory pain, the mechanisms driving its sensitisation during inflammation remain poorly understood. Here, we found that an inflammatory soup of bradykinin, interleukin 1{beta} (IL-1{beta}) and tumour necrosis factor (TNF) sensitised TRPM3 function in isolated mouse sensory neurons; IL-1{beta} and TNF, but not bradykinin, independently potentiated TRPM3 function. TRPM3 expression and translocation to the membrane remained unchanged upon individual or combined exposure to these inflammatory mediators, which suggests post-translational modification occurs. Finally, using the model of complete Freund\'s adjuvant-induced knee inflammation, we found that pharmacological blockade of TRPM3 does not alleviate inflammatory pain, which contrasts with previous reports using different pain models. We propose that the nuances of the immune response may determine the relative contribution of TRPM3 to nociceptive signalling in different neuro-immune contexts. Collectively, our findings improve insight into the role of TRPM3 sensitisation in inflammatory pain. | 2024-09-14 | |
Operation of spinal sensorimotor circuits controlling phase durations during tied-belt and split-belt locomotion after a lateral thoracic hemisection | 10.1101/2024.09.10.612376 | Rybak, I. A.; Shevtsova, N. A.; Audet, J.; Yassine, S.; Markin, S. N.; Prilutsky, B. I.; Frigon, A. | Locomotion is controlled by spinal circuits that interact with supraspinal drives and sensory feedback from the limbs. These sensorimotor interactions are disrupted following spinal cord injury. The thoracic lateral hemisection represents an experimental model of an incomplete spinal cord injury, where connections between the brain and spinal cord are abolished on one side of the cord. To investigate the effects of such an injury on the operation of the spinal locomotor network, we used our computational model of cat locomotion recently published in eLife (Rybak et al., 2024) to investigate and predict changes in cycle and phase durations following a thoracic lateral hemisection during treadmill locomotion in tied-belt (equal left-right speeds) and split-belt (unequal left-right speeds) conditions. In our simulations, the hemisection was always applied to the right side. Based on our model, we hypothesized that following hemisection, the contralesional (intact, left) side of the spinal network is mostly controlled by supraspinal drives, whereas the ipsilesional (hemisected, right) side is mostly controlled by somatosensory feedback. We then compared the simulated results with those obtained during experiments in adult cats before and after a mid-thoracic lateral hemisection on the right side in the same locomotor conditions. Our experimental results confirmed many effects of hemisection on cat locomotion predicted by our simulations. We show that having the ipsilesional hindlimb step on the slow belt, but not the fast belt, during split-belt locomotion substantially reduces the effects of lateral hemisection. The model provides explanations for changes in temporal characteristics of hindlimb locomotion following hemisection based on altered interactions between spinal circuits, supraspinal drives, and somatosensory feedback. | 2024-09-14 | |
Soil redox drives virus-host community dynamics and plant biomass degradation in tropical rainforest soils | 10.1101/2024.09.13.612973 | Trubl, G.; Leleiwi, I.; Campbell, A.; Kimbrel, J. A.; Bhattacharyya, A.; Riley, R.; Malmstrom, R. R.; Blazewicz, S. J.; Pett-Ridge, J. | Background: Wet tropical forest soils store a vast amount of organic carbon and cycle over a third of terrestrial net primary production. The microbiomes of these soils have a global impact on greenhouse gases and tolerate a remarkably dynamic redox environment-driven by high availability of reductant, high soil moisture, and fine-textured soils that limit oxygen diffusion. Yet tropical soil microbiomes, particularly virus-host interactions, remain poorly characterized, and we have little understanding of how they will shape future soil carbon cycling as high-intensity drought and precipitation events make soil redox conditions less predictable. Results: To investigate the effects of shifting soil redox conditions on active viral communities and virus-microbe interactions, we conducted a 44-day redox manipulation experiment using soils from the Luquillo Experimental Forest, Puerto Rico, amended with 13C-enriched plant biomass. We sequenced 10 bulk metagenomes and 85 stable isotope probing targeted metagenomes generated by extracting whole community DNA, performing density fractionation, and conducting shotgun sequencing. Viral and microbial genomes were assembled resulting in 5,420 viral populations (vOTUs) and 927 medium-to-high-quality metagenome-assembled genomes across 25 bacterial phyla. Notably, over half (54%) of the vOTUs were 13C-enriched, highlighting their active role in microbial degradation of plant litter. These active vOTUs primarily infected bacterial phyla Pseudomonadota, Acidobacteriota, and Actinomycetota, and 57% were unique to a particular redox treatment. The anoxic samples exhibited the most distinct viral communities, with an increased potential for modulating host metabolism by carrying redox-specific glycoside hydrolases. However, over a third of the vOTUs were present in all redox conditions, suggesting selection for cosmopolitan viruses occurs in these soils that naturally experience dynamic redox conditions. Conclusions: Our study demonstrates how redox conditions shape viral communities and virus-host interactions in soils. By applying different DNA assembly methods on stable isotope probing targeted metagenomes and incubating soils under various redox regimes, we identified distinct viral populations and observed significant variations in viral community composition and function. These findings highlight the specialized roles of viruses in microbial carbon degradation under diverse environmental conditions, providing important insights into their contributions to carbon cycling and the broader implications for climate change. | 2024-09-14 |