Biology subjects

He, H.

Publications and source records attributed to He, H..

At least 91 records · Page 5Linked to original sources

bioRxiv2024

Non-invasive optoacoustic imaging visualizes exercise-induced dermal revascularization in obese mice

Microcirculatory dysfunction in dermal (dWAT) and subcutaneous white adipose tissue (scWAT) of obese humans may predict cardio-metabolic disease progression. In-vivo visualization and monitoring of microvascular remodeling in these tissues remains challenging. We compared performance of multi-spectral optoacoustic tomography (MSOT) and raster-scanning optoacoustic mesoscopy (RSOM) in visualizing lipid and hemoglobin contrast in scWAT and dWAT of diet-induced obese (DIO) mice undergoing voluntary wheel running. MSOT quantitatively visualized lipid and hemoglobin contrast in fat depots at early stages of DIO. RSOM precisely visualizes microvasculature with quantitative readouts of skin layer thickness and vascular density in dWAT and dermis. Combination of MSOT and RSOM resolved exercise-induced morphological changes in microvasculature density, tissue oxygen saturation, lipid and blood volume content in dWAT and scWAT. Combination of MSOT and RSOM precisely monitor microcirculatory dysfunction and intervention response in dWAT and scWAT of DIO mice. Our findings lay out the foundation for future clinical studies using optoacoustic-derived vascular readouts from adipose tissues as a biomarker for monitoring microcirculatory function in cardio-metabolic disease.

bioengineering↗

bioRxiv2024

Click-free imaging of carbohydrate trafficking in live cells using an azido photothermal probe

Real-time tracking of intracellular carbohydrates remains challenging. While click chemistry allows bio-orthogonal tagging with fluorescent probes, the reaction permanently alters the target molecule and only allows a single snapshot. Here, we demonstrate click-free mid-infrared photothermal (MIP) imaging of azide-tagged carbohydrates in live cells. Leveraging the micromolar detection sensitivity for 6-azido-trehalose (TreAz) and the 300-nm spatial resolution of MIP imaging, the trehalose recycling pathway in single mycobacteria, from cytoplasmic uptake to membrane localization, is directly visualized. A peak shift of azide in MIP spectrum further uncovers interactions between TreAz and intracellular protein. MIP mapping of unreacted azide after click reaction reveals click chemistry heterogeneity within a bacterium. Broader applications of azido photothermal probes to visualize the initial steps of the Leloir pathway in yeasts and the newly synthesized glycans in mammalian cells are demonstrated.

cell biology↗

bioRxiv2024

Exo70 promotes herpesvirus secretory vesicle tethering and virion release in an exocyst complex-dependent manner

The exocyst complex, a multisubunit protein complex, mediates secretory vesicles fusion with the plasma membrane (PM) to deliver materials to the cell surface or to release cargoes to the extracellular space, but whether it is related to viral secretory vesicles tethering and virion release is unknown. Here, we identified that the exocyst complex subunit Exo70 promotes the trafficking of herpesvirus secretory vesicles and the release of mature virions in an exocyst complex-dependent manner. Mechanistically, mutation of Exo70 Lys632 and Lys635 diminishes viral secretory vesicles anchoring to PM. Moreover, the small GTPase Rab11a is necessary for the transport of viral secretory vesicles, and the Snapin-Exo70-SNAP23 axis is involved in fusion of viral secretory vesicles to the PM and release of virions. Most significantly, we discovered that Endosidin2 (ES2), an inhibitor of exocytosis via the exocyst complex, provides protection against herpesvirus infection in cells and mice. Overall, these findings unveil a previously uncharacterized role and mechanism of the exocyst complex in viral replication, highlighting its potential as an effective strategy against herpesvirus infection. Author summaryHerpesviruses, such as herpes simplex virus 1 (HSV-1) and bovine herpes virus type 1 (BoHV-1), are highly prevalent pathogens that establish lifelong infections and cause various diseases in humans and animals. While the mechanisms underlying the transport of herpesvirus secretory vesicles from the Golgi to the PM and the subsequent viral release are still poorly understood. In our study, we have discovered that Exo70 promotes the trafficking of HSV-1 and BoHV-1 secretory vesicles and facilitates the release of mature virions in an exocyst complex-dependent manner. Mechanistically, both knockdown of Exo70 and a phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2)-binding-deficient mutant of Exo70 caused virions to be trapped in the cytoplasm and unable to tether to PM, indicating that the binding of Exo70 to PI(4,5)P2 is critical for the docking virus secretory vesicles with PM. Additionally, the Snapin-Exo70-SNAP23 axis plays a pivotal role in the fusion between viral secretory vesicles and the PM, ultimately facilitating the release of virions. We found that Rab11a is necessary for viral secretory vesicle trafficking mediated by Exo70. Most importantly, we discovered that ES2, a transport inhibitor that combines with Exo70 to terminate the final stage of exocytosis via the exocyst complex, can protect cells and mice from -herpesvirus infection.

microbiology↗

bioRxiv2024

Structural insights into sphingosine 1-phosphate receptor 4 activation

S1PR4 is one of five subtypes of sphingosine 1-phosphate receptors (S1PRs) that regulate immune cell functioning, with functional distinctions to other subtypes. S1PR1-targeted modulators caused serious cardiac and vascular adverse effects because S1PR1 was expressed throughout the whole body. Since S1PR4 was only expressed in lung and lymphoid cells, S1PR4-targeted modulators might not trigger these side effects. However, the development of S1PR4-specific agonists is greatly hindered because of the lack of activated S1PR4 structure. Here, we resolved cryo-EM structures of activated S1PR4 and revealed the structural mechanism of ligand recognition, receptor activation, and Gi coupling. Our results offered structural templates for the development of selective S1PR4 agonists with improved safety profiles.

biochemistry↗

bioRxiv2023

PRDM3/16 Regulate Chromatin Accessibility Required for NKX2-1 Mediated Alveolar Epithelial Differentiation and Function

Differential chromatin accessibility accompanies and mediates transcriptional control of diverse cell fates and their differentiation during embryogenesis. While the critical role of NKX2-1 and its transcriptional targets in lung morphogenesis and pulmonary epithelial cell differentiation is increasingly known, mechanisms by which chromatin accessibility alters the epigenetic landscape and how NKX2-1 interacts with other co-activators required for alveolar epithelial cell differentiation and function are not well understood. Here, we demonstrate that the paired domain zinc finger transcriptional regulators PRDM3 and PRDM16 regulate chromatin accessibility to mediate cell differentiation decisions during lung morphogenesis. Combined deletion of Prdm3 and Prdm16 in early lung endoderm caused perinatal lethality due to respiratory failure from loss of AT2 cell function. Prdm3/16 deletion led to the accumulation of partially differentiated AT1 cells and loss of AT2 cells. Combination of single cell RNA-seq, bulk ATAC-seq, and CUT&RUN demonstrated that PRDM3 and PRDM16 enhanced chromatin accessibility at NKX2-1 transcriptional targets in peripheral epithelial cells, all three factors binding together at a multitude of cell-type specific cis-active DNA elements. Network analysis demonstrated that PRDM3/16 regulated genes critical for perinatal AT2 cell differentiation, surfactant homeostasis, and innate host defense. Lineage specific deletion of PRDM3/16 in AT2 cells led to lineage infidelity, with PRDM3/16 null cells acquiring partial AT1 fate. Together, these data demonstrate that NKX2-1-dependent regulation of alveolar epithelial cell differentiation is mediated by epigenomic modulation via PRDM3/16. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=130 HEIGHT=200 SRC="FIGDIR/small/570481v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@500f6org.highwire.dtl.DTLVardef@821daaorg.highwire.dtl.DTLVardef@69ba87org.highwire.dtl.DTLVardef@154f754_HPS_FORMAT_FIGEXP M_FIG C_FIG Model of the role of PRMD3/16 in alveolar developmentPRMD3/16 participate in cell fate specification in the lung by modulating chromatin accessibility (top row) and by partnering with NKX2-1 and partner transcription factors to drive gene expression (second row) via a gene regulatory network required for terminal cell differentiation and surfactant expression in AT2 cells (third row). Loss of PRDM3/16 activity in lung endoderm leads to reduced AT2 quorum, failure of AT2 surfactant function, and transition to an immature AT1 phenotype (bottom panel).

developmental biology↗

bioRxiv2023

ZNF397 Loss Triggers TET2-driven Epigenetic Rewiring, Lineage Plasticity, and AR-targeted Therapy Resistance in AR-dependent Cancers

Cancer cells exhibit phenotypical plasticity and epigenetic reprogramming, which allows them to evade lineage-dependent targeted treatments by adopting lineage plasticity. The underlying mechanisms by which cancer cells exploit the epigenetic regulatory machinery to acquire lineage plasticity and therapy resistance remain poorly understood. We identified Zinc Finger Protein 397 (ZNF397) as a bona fide co-activator of the androgen receptor (AR), essential for the transcriptional program governing AR-driven luminal lineage. ZNF397 deficiency facilitates the transition of cancer cell from an AR-driven luminal lineage to a Ten-Eleven Translocation 2 (TET2)-driven lineage plastic state, ultimately promoting resistance to therapies inhibiting AR signaling. Intriguingly, our findings indicate that TET2 inhibitor can eliminate the AR targeted therapies resistance in ZNF397-deficient tumors. These insights uncover a novel mechanism through which prostate and breast cancers acquire lineage plasticity via epigenetic rewiring and offer promising implications for clinical interventions designed to overcome therapy resistance dictated by lineage plasticity. Statement of SignificanceThis study reveals a novel epigenetic mechanism regulating tumor lineage plasticity and therapy response, enhances understanding of drug resistance and unveils a new therapeutic strategy for prostate cancer and other malignancies. Our findings also illuminate TET2s oncogenic role and mechanistically connect TET2-driven epigenetic rewiring to lineage plasticity and therapy resistance.

cancer biology↗

bioRxiv2023

Systems genetics identifies the association between Enc1 and cognitive function in the hippocampus

Ectodermal-Neural Cortex 1 (ENC1) is expressed in multiple regions of the brain, including the hippocampus. However, knowledge about its function has been well explored only in the context of peroxidative stress and cancer. In this study, we investigated the association of hippocampal Enc1 with cognitive function in BXD mice. We performed Pearson correlation, phenotype-wide association analysis (PheWAS), expression-Based PheWAS, pathway enrichment, and protein interaction networks on Enc1 and BXD phenotypes/transcriptome of the hippocampus, and the results indicated that Enc1 is inextricably linked to cognitive performance. In addition, we found that most of the Enc1 co-expressed genes were highly expressed in GABAergic neuronal cells. Expression quantitative trait loci analysis indicated that Enc1 was cis-regulated in the hippocampus of mice as well as human. Genome-wide association analysis revealed ENC1 to be significantly associated with cognitive-related traits, including age-related cognitive changes etc. In conclusion, our findings demonstrated that Enc1 is involved in cognitive functions mainly in hippocampal GABAergic neuronal cells through neurogenesis, synaptic signaling, and CGMP-PKG signaling pathways, and interacts with the neurological function-related genes.

genetics↗

bioRxiv2023

De novo generation of antibody CDRH3 with a pre-trained generative large language model

Artificial Intelligence (AI) techniques have made great advances in assisting antibody design. However, antibody design still heavily relies on isolating antigen-specific antibodies from serum, which is a resource-intensive and time-consuming process. To address this issue, we propose a Pre-trained Antibody generative large Language Model (PALM) for the de novo generation of artificial antibodies heavy chain complementarity-determining region 3 (CDRH3) with desired antigen-binding specificity, reducing the reliance on natural antibodies. We also build a high-precision model antigen-antibody binder (A2binder) that pairs antigen epitope sequences with antibody sequences to predict binding specificity and affinity. PALM-generated antibodies exhibit binding ability to SARS-CoV-2 antigens, including the emerging XBB variant, as confirmed through in-silico analysis and in-vitro assays. The in-vitro assays validated that PALM-generated antibodies achieve high binding affinity and potent neutralization capability against both wild-type and XBB spike proteins of SARS-CoV-2. Meanwhile, A2binder demonstrated exceptional predictive performance on binding specificity for various epitopes and variants. Furthermore, by incorporating the attention mechanism into the PALM model, we have improved its interpretability, providing crucial insights into the fundamental principles of antibody design.

bioinformatics↗

bioRxiv2023

SACNet: A Multiscale Diffeomorphic Convolutional Registration Network with Prior Neuroanatomical Constraints for Flexible Susceptibility Artifact Correction in Echo Planar Imaging

Susceptibility artifacts (SAs), inevitable in brain diffusion MR (dMRI) scans acquired using single-shot echo planar imaging (EPI), severely compromise the accurate detection of human brain structure. Existing SA correction (SAC) methods offer inadequate correction quality and limited applicability across diverse datasets with varied acquisition protocols. To address these challenges, we proposed SACNet, a SAC framework based on unsupervised registration convolutional networks, featuring: i) a novel diffeomorphism regularization function to avoid unnatural SAC warps, modified from a potential well function; ii) an integration with prior neuroanatomical constraints and coarse-to-fine processing strategy to enables multi-scale geometric and intensity recoveries in severe distorted areas; iii) a unified registration framework that incorporates multiple phase-encoding (PE) EPI images and structural images, ensuring compatibility with both single- and inverse-PE protocols, with or without field maps. Utilizing simulated dMRI images and over 2000 brain scans from neonatal, child, adult and traveling participants, our method consistently demonstrates state-of-the-art correction performance. Notably, SACNet effectively reduces SAs-related multicenter effects compared to existing methods. We have developed user-friendly tools using containerization techniques, hope to facilitate SAC correction quality across extensive neuroimaging studies.

neuroscience↗

bioRxiv2023

Local two-photon excitation reveals the mechanism of mitochondrial Ca2+ release

Mitochondrial Ca2+ (mitoCa2+) simultaneously implicates respiration, mitochondrial physiology, and cell signaling, which prevents the disentanglement of mitoCa2+ from those complex processes. Although mitochondria have long been recognized as temporary Ca2+ buffer, how mitoCa2+ is regulated and released remains unclear. In this study, we report a specific photochemical excitation to flavoproteins in single-mitochondrion tubulars by a tightly-focused femtosecond laser that triggers local mitoCa2+ transients, without any extra-mitochondrial Ca2+ involved. The two-photon excitation to complex I and II accelerates the entire electron transport chain (ETC) and promotes ATP synthesis. Free mitoCa2+ is thus released from the Ca2+-phosphate ion (Pi) complexes at complex V in mitochondrial matrix during ATP synthesis to form mitoCa2+ transients there. The abnormal mitoCa2+ signaling by knockdown of ATP synthase subunit affects cell proliferation, apoptosis, and mitophagy. Our results reveal mitoCa2+ is released and regulated by ETC and ATP synthesis rather than the reverse. Mitochondria are multi-functional units to simultaneously produce energy and coordinate molecular signaling for cells. The key process of metabolism, tricarboxylic acid (TCA) cycle, is continuously ongoing in mitochondrial matrix and generates reducing equivalents for subsequent electron transfer (Martinez-Reyes and Chandel, 2020; Martinez-Reyes et al., 2016). The respiration is accomplished by electron transport chain (ETC) in the inner mitochondrial membrane (IMM) (Marreiros et al., 2016). Mitochondria regulate a series of signaling cascades to mediate apoptosis, autophagy, and cell senescence (Galluzzi et al., 2014; Rasola and Bernardi, 2011; Ziegler et al., 2021). Biosynthesis of some amino acids and nucleotides also takes place in mitochondria (Ahn and Metallo, 2015; Li and Hoppe, 2023). Those processes are involved with each other through complex crosstalk and feedbacks. Mitochondria have long been recognized as Ca2+ buffer to temporarily deposit abnormal cytosolic Ca2+ for cellular Ca2+ homeostasis (Garbincius and Elrod, 2022; Kirichok et al., 2004; Lambert et al., 2019). However, the Ca2+ entry into mitochondria definitely influence those mitochondrial processes and functions (Garbincius and Elrod, 2022). The acute and direct consequences of Ca2+ entry into mitochondria are found as depolarization of mitochondrial membrane potential (MMP). If the cellular Ca2+ level is too high, mitochondria may become damaged and dysfunctional. Mitophagy/autophagy and even apoptosis are initiated (Galluzzi et al., 2014; Lou et al., 2020; Rasola and Bernardi, 2011). Moreover, Ca2+ in mitochondrial matrix (mitoCa2+) takes an essential role in quite a few physiological processes including mitochondrial fission and fusion, cell development, and proliferation (Doonan et al., 2014; Singh and Mabalirajan, 2021; Steffen and Koehler, 2018). The respiration is also believed to be related to mitoCa2+ (Glancy and Balaban, 2012; Wescott et al., 2019). Technically, perturbation to mitoCa2+ affects most mitochondrial processes and physiology, which makes it quite difficult to interrogate how Ca2+ is regulated and released in mitochondria. So far, the regulation of mitoCa2+ remains in mist. In this study, we report a single-mitochondrion photochemical process by tightly-focused femtosecond laser that specifically excites complex I and II by two-photon excitation and accelerate ETC. Free mitoCa2+ is released from Ca2+-phosphate ion (Pi) complexes by ATP synthesis at complex V in mitochondria. These results clarify the mechanism of mitoCa2+ regulation and provide further insights in the relationship between mitoCa2+ and respiration.

cell biology↗

bioRxiv2023

Cooperative progression of colitis and leukemia modulated by clonal hematopoiesis via PTX3/IL-1β pro-inflammatory signaling

Clonal hematopoiesis (CH) is considered an important risk factor for all-cause mortality and the development of multiple chronic diseases including hematological neoplasms, cardiovascular diseases, and potentially a range of autoimmune or immune-deficiency diseases. Mutations in TET2 are one of the first identified, most important, and prevalent genetic drivers of CH. However, cooperative factors and mechanisms underlying TET2-deficiency related CH (TedCH) remain largely unknown. Recently, it has been suggested that certain diseases occurred before TedCH and promote TedCH trajectory on the contrary, indicating that diseases in non-hematopoietic organs may act as environmental non-genetic drivers of CH. To clarify the relationships between immune-dysfunctional diseases and CH, here we tested the impact of various challenges on TedCH. We found that expedited TedCH depended on establishment of an inflammatory environment. Primary or chimeric Tet2-mutant mice spontaneously developed co-symptoms reminiscent of human chronic colitis and myeloid leukemia, which was exacerbated by feeding with DSS, an experimental inducer of ulcerative colitis. Single cell RNA-seq (scRNA-seq) analysis reveals in depth the damage of colon in the Tet2-mutant mice in physiological conditions or fed with DSS, along with increase of dysbacteriosis indicated by gut microbiome analysis. Results from colon scRNA-seq from both mouse and human highlight the important roles of PTX3/IL-1{beta} pro-inflammatory signaling in promoting colitis or leukemia. Finally, TedCH trajectory and inflammation in colon and bone marrow were ameliorated by treatment of IL-1R1 inhibitor Anakinra. Our study suggests that PTX3/IL-1{beta} signaling and clonal hematopoiesis cooperate and play important roles in gut-bone marrow axis and related diseases including colitis and leukemia. HighlightsO_LICertain environmental factors, such as Dextran Sulfate Sodium (DSS), an experimental inducer of ulcerative colitis, promote TedCH C_LIO_LIColitis and leukemia are spontaneously and simultaneously developed in Tet2-defficient primary or chimeric mice, along with increased pathogenic gut microbiomes, indicating an aberrant gut-bone marrow axis in the mutant mice. C_LIO_LISingle cell RNA-seq analysis reveals enhanced PTX3, a soluble pattern recognition molecule and IL-1{beta} pro-inflammatory signaling in colitis and leukemia. C_LIO_LIThe In vivo function of the PTX3/IL-1{beta} pro-inflammatory signaling in TedCH is indicated in human colitis and validated in experimental settings. C_LI

immunology↗

bioRxiv2023

Water and chloride as allosteric inhibitors in WNK kinase osmosensing

Osmotic stress and chloride regulate the autophosphorylation and activity of the WNK1 and WNK3 kinase domains. The kinase domain of unphosphorylated WNK1 (uWNK1) is an asymmetric dimer possessing water molecules conserved in multiple uWNK1 crystal structures. Conserved waters are present in two networks, referred to here as conserved water networks 1 and 2 (CWN1and CWN2). Here we show that PEG400 applied to crystals of dimeric uWNK1 induces de-dimerization. Both the WNK1 the water networks and the chloride binding site and are disrupted by PEG400. CWN1 is surrounded by a cluster of pan-WNK-conserved charged residues. Here we mutagenized these charges in WNK3, a highly active WNK isoform kinase domain, and WNK1, the isoform best studied crystallographically. Mutation of E314 in the Activation Loop of WNK3 (WNK3/E314Q and WNK3/E314A, and the homologous WNK1/E388A) enhanced the rate of autophosphorylation, and reduced chloride sensitivity. Other WNK3 Cluster mutants reduced the rate of autophosphorylation activity coupled with greater chloride sensitivity than wild-type. The water and chloride regulation thus appear linked. The lower activity of some mutants may reflect effects on catalysis. Crystallography showed that activating mutants introduced conformational changes in similar parts of the structure to those induced by PEG400. WNK activating mutations and crystallography support a role for CWN1 in WNK inhibition consistent with water functioning as an allosteric ligand.

biophysics↗

bioRxiv2023

NRF2 Regulates Contractile Vascular Smooth Muscle Cells Phenotypic Switching In Abdominal Aortic Aneurysm

OBJECTIVEEvidence from previous studies has demonstrated that NRF2 can protect cardiovascular progression,which prompted us to study if NRF2 is involved in AAA development. APPROACH AND RESULTSThe difference and function of NRF2 in AAA were verified on human aortic tissues combined with scRNA-seq samples.We then used angiotensin II (Ang II) infusion mouse model with VSMCs-specific knockout to study the role of NRF2 in AAA formation.Primary cultured VSMCs were used to study how NRF2 regulates contractile VSMCs phenotype.Patient specimens were obtained to investigate the relevance of NRF2 expression to human AAA disease. NRF2 was induced in abdominal aortic VSMCs in both mouse and human AAA tissues. VSMCs-specific NRF2 knockout increased experimental AAA formation. Mechanistically, NRF2 regulates the contractile VSMCs phenotypic through miRNA145, leading to increased classic contractile protein level to prevent the development of AAA. CONCLUSIONSNRF2 is a negative regulator of AAA development, and thus may represent a potentially new therapeutic target to inhibit AAA growth and rupture.

bioinformatics↗

bioRxiv2023

YiNet: An Integrated Traditional Chinese and Western Medicine Platform for Viral Infectious Diseases

Viral infectious diseases (VIDs) impose a heavy burden on global public health. Traditional Chinese medicine (TCM) has previously and is currently contributing to the prevention and treatment of infectious diseases. Omics and information technology enable the precise identification of virus characteristics, virus-host interactions and TCM mechanisms. In this study, we constructed the YiNet platform to better integrate the novel techniques and historical experience of TCM in infectious diseases. YiNet comprises three modules: knowledge base, database and toolkit. The YiNet knowledge base involves 43 VIDs, thereby systematically integrating the knowledge regarding viruses, host symptoms and TCM and Western medicine (commonly used chemical drugs, 6,899 herbs and 2,481 formulas). The YiNet database module includes multiple databases, comprising 57,340 genome sequences of 45,791 viral strains and 5,726 multi-omics datasets such as RNA-seq, ChIP-seq and ATAC-seq from different tissues and cell models. It also integrates 1,105 real-world TCM clinical cases. We adopted visual analysis tool to investigate pathogen-host-herb relationships. To explore pharmacological mechanisms for the core herbs, we added formula data mining and network pharmacology analysis pipelines and visualisation tools. YiNet can facilitate the mechanistic study of TCM and drug development for VIDs. The YiNet platform is publicly available at http://yinet.gene.ac/.

bioinformatics↗

bioRxiv2023

GenomeMUSter mouse genetic variation service enables multi-trait, multi-population data integration and analyses

Hundreds of inbred laboratory mouse strains and intercross populations have been used to functionalize genetic variants that contribute to disease. Thousands of disease relevant traits have been characterized in mice and made publicly available. New strains and populations including the Collaborative Cross, expanded BXD and inbred wild-derived strains add to set of complex disease mouse models, genetic mapping resources and sensitized backgrounds against which to evaluate engineered mutations. The genome sequences of many inbred strains, along with dense genotypes from others could allow integrated analysis of trait - variant associations across populations, but these analyses are not feasible due to the sparsity of genotypes available. Moreover, the data are not readily interoperable with other resources. To address these limitations, we created a uniformly dense data resource by harmonizing multiple variant datasets. Missing genotypes were imputed using the Viterbi algorithm with a data-driven technique that incorporates local phylogenetic information, an approach that is extensible to other model organism species. The result is a web- and programmatically-accessible data service called GenomeMUSter (https://muster.jax.org), comprising allelic data covering 657 strains at 106.8M segregating sites. Interoperation with phenotype databases, analytic tools and other resources enable a wealth of applications including multi-trait, multi-population meta-analysis. We demonstrate this in a cross-species comparison of the meta-analysis of Type 2 Diabetes and of substance use disorders, resulting in the more specific characterization of the role of human variant effects in light of mouse phenotype data. Other applications include refinement of mapped loci and prioritization of strain backgrounds for disease modeling to further unlock extant mouse diversity for genetic and genomic studies in health and disease.

genetics↗

bioRxiv2023

Heterogeneity and Functional Analysis of Cardiac Fibroblasts in Heart Development

BackgroundAs one of the major cell types in the heart, fibroblasts play critical roles in multiple biological processes. Cardiac fibroblasts are known to develop from multiple sources, but their transcriptional profiles have not been systematically compared. Furthermore, while the function of a few genes in cardiac fibroblasts has been studied, the overall function of fibroblasts as a cell type remains uninvestigated. MethodsSingle-cell mRNA sequencing (scRNA-seq) and bioinformatics approaches were used to analyze the genome-wide genes expression and extracellular matrix genes expression in fibroblasts, as well as the ligand-receptor interactions between fibroblasts and cardiomyocytes. Single molecular in situ hybridization was employed to analyze the expression pattern of fibroblast subpopulation-specific genes. The Diphtheria toxin fragment A (DTA) system was utilized to ablate fibroblasts at each developmental phase. ResultsUsing RNA staining of Col1a1 at different stages, we grouped cardiac fibroblasts into four developmental phases. Through the analysis of scRNA-seq profiles of fibroblasts at 18 stages from two mouse strains, we identified significant heterogeneity, preserving lineage gene expression in their precursor cells. Within the main fibroblast population, we found differential expressions of Wt1, Tbx18, and Aldh1a2 genes in various cell clusters. Lineage tracing studies showed Wt1- and Tbx18-positive fibroblasts originated from respective epicardial cells. Furthermore, using a conditional DTA system-based elimination, we identified the crucial role of fibroblasts in early embryonic and heart growth, but not in neonatal heart growth. Additionally, we identified the zone- and stage-associated expression of extracellular matrix genes and fibroblast-cardiomyocyte ligand-receptor interactions. This comprehensive understanding sheds light on fibroblast function in heart development. ConclusionWe observed cardiac fibroblast heterogeneity at embryonic and neonatal stages, with preserved lineage gene expression. Ablation studies revealed their distinct roles during development, likely influenced by varying extracellular matrix genes and ligand-receptor interactions at different stages.

developmental biology↗

bioRxiv2023

Inflammation plays a critical role in damage to the bronchiolar epithelium induced by Trueperella pyogenes in vitro and in vivo

Trueperella pyogenes can cause severe pulmonary disease in swine, but the mechanism of pathogenesis is not well defined. T. pyogenes-induced damage to porcine bronchial epithelial cells (PBECs), porcine precision-cut lung slices (PCLS) and respiratory epithelium of mice remains unknown. In this study, we used T. pyogenes 20121 to infect PBECs in air-liquid interface conditions and porcine PCLS. T. pyogenes could adhere to, colonize and induce cytotoxic effect on PBECs and the luminal surface of bronchi in PCLS, which damaged the bronchiolar epithelium. Moreover, bronchiolar epithelial cells showed extensive degeneration in infected mice lungs. Furthermore, western blot showed the NOD-like receptor (NLR)/ C-terminal caspase recruitment domain (ASC)/caspase-1 axis and nuclear factor-kappa B (NF-{kappa}B) pathway were involved in inflammation in PCLS and lungs of mice, which also confirms PCLS provide a platform to analyze pulmonary immune response. Meanwhile, the levels of p-c-Jun N-terminal kinase (JNK), p-extracellular signal-regulated kinase (ERK) and p-protein kinase B (AKT) were increased significantly, which indicated the mitogen-activated protein kinase (MAPK) and Akt pathways were also involved of inflammation in T. pyogenes-infected mice. In addition, we used T. pyogenes 20121 to infect tumour necrosis factor alpha (TNF-)-/- mice, the results indicated apoptosis and injury in respiratory epithelium of infected TNF--/- mice were alleviated. Thus, pro-inflammatory cytokine TNF- played a role in apoptosis and respiratory epithelium injury of mice lungs. Collectively, our study provides an insight into the inflammatory injury induced by T. pyogenes, and suggests that blocking NLR or TNF- may be a potential therapeutic strategy against T. pyogenes infection.

immunology↗

bioRxiv2023

Maresin-2 Fine-tunes ULK1 O-GlcNAcylation to Improve Post Myocardial Infarction Remodeling

Maresin-2, a specialized pro-solving mediator of inflammation has been consolidated to be a novel cytokine fine-tuning inflammatory cascade. However, the underlying molecular basis is still largely unknown. Focused on cardiac dysfunction and remodeling, we employed in vivo- and in vitro- based genome editing methodology tools including adenosine associated virus, adenosine virus, lenti-virus, plasmid transfection, and CRISPR-Cas9 methodology for investigation. As suggested, exogenous maresin-2 supplement facilitated autophagosome formation by microtubule-associated proteins 1A/1B light chain 3B (LC3) conjugation system under the modulation of O-GlcNAcylation dependent ULK1 activation, whereas reversed by ULK1 S409A and S422A mutagenesis, showcasing the potential O-GlcNAc (O-linked {beta}- N-acetylglucosamine) modifiable sites on ULK1. Moreover, we found that hereafter maresin-2 treatment glutamine-fructose-6-phosphate aminotransferase 1 (GFAT1) which is accessary to sense hexosamine biosynthesis influx is more likely the prime checkpoint for conjugating O-terminal {beta}-N-acetylglucosamine motif onto ULK1, rather than O-linked N-acetylglucosaminyltransferase (OGT). Mechanistically, maresin-2 largely prohibits transforming growth factor-{beta} (TGF-{beta})-activated kinase 1 (TAK1), therefore increasing the availability of TAB1 for GFAT1, which encourages O-GlcNAcylation of ULK1.

molecular biology↗