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Gao, H.

Publications and source records attributed to Gao, H..

At least 145 records · Page 8Linked to original sources
bioRxiv2022

Development of highly potent non-covalent inhibitors of SARS-CoV-2 3CLpro

The SARS-CoV-2 virus is the causal agent of the ongoing pandemic of coronavirus disease 2019 (COVID-19). There is an urgent need for potent, specific antiviral compounds against SARS-CoV-2. The 3C-like protease (3CLpro) is an essential enzyme for the replication of SARS-CoV-2 and other coronaviruses, and thus is a target for coronavirus drug discovery. Nearly all inhibitors of coronavirus 3CLpro reported so far are covalent inhibitors. Here, we report the development of specific, non-covalent inhibitors of 3CLpro. The most potent one, WU-04, effectively blocks SARS-CoV-2 replications in human cells with EC50 values in the 10-nM range. WU-04 also inhibits the 3CLpro of SARS-CoV and MERS-CoV with high potency, indicating that it is a pan-inhibitor of coronavirus 3CLpro. WU-04 showed anti-SARS-CoV-2 activity similar to that of PF-07321332 (Nirmatrelvir) in K18-hACE2 mice when the same dose was administered orally. Thus, WU-04 is a promising drug candidate for coronavirus treatment. One-Sentence SummaryA oral non-covalent inhibitor of 3C-like protease effectively inhibits SARS-CoV-2 replication.

microbiology↗
bioRxiv2022

Origin, Prospective Identification, and Function of Circulating Endothelial Colony Forming Cells in Mouse and Human

Most circulating endothelial cells are apoptotic, but rare circulating endothelial colony forming cells (C-ECFCs) with proliferative and vasculogenic activity can be cultured; the origin and naive function of these C-ECFCs remains obscure. Herein, detailed lineage tracing reveals murine C-ECFCs emerge in the early postnatal period, display high vasculogenic potential, with enriched frequency of clonal proliferative cells compared to tissue-resident ECFCs, and are not committed to or derived from the bone marrow hematopoietic system but from tissue-resident ECFCs. In human subjects, C-ECFCs are present in the CD34bright cord blood mononuclear subset, possess proliferative potential and in vivo vasculogenic function in a naive or cultured state, and display a single cell transcriptome sharing some umbilical venous endothelial cell features like, higher Protein C Receptor and extracellular matrix gene expression. This study provides an advance for the field by identifying the origin, naive function, and antigens to prospectively isolate C-ECFCs for translational studies.

cell biology↗
bioRxiv2022

Single Cell Cortical Bone Transcriptomics Defines Novel Osteolineage Gene Sets Altered in Chronic Kidney Disease

Due to a lack of spatial-temporal resolution at the single cell level, the etiologies of the bone dysfunction caused by diseases such as normal aging, osteoporosis, and the metabolic bone disease associated with chronic kidney disease (CKD) remain largely unknown. To this end, flow cytometry and scRNAseq were performed on long bone cells from Sost-cre/Ai9+ mice, and pure osteolineage transcriptomes were identified, including novel osteocyte-specific gene sets. Clustering analysis isolated osteoblast precursors that expressed Tnc, Mmp13, and Spp1, and a mature osteoblast population defined by Smpd3, Col1a1, and Col11a1. Osteocytes were demarcated by Cd109, Ptprz1, Ramp1, Bambi, Adamts14, Spns2, Bmp2, WasI, and Phex. We validated our in vivo scRNAseq using integrative in vitro promoter occupancy via ATACseq coupled with transcriptomic analyses of a conditional, temporally differentiated MSC cell line. Further, trajectory analyses predicted osteoblast-to-osteocyte transitions via defined pathways associated with a distinct metabolic shift as determined by single-cell flux estimation analysis (scFEA). Using the adenine mouse model of CKD, at a time point prior to major skeletal alterations, we found that gene expression within all stages of the osteolineage was disturbed. In sum, distinct populations of osteoblasts/osteocytes were defined at the single cell level. Using this roadmap of gene assembly, we demonstrated unrealized molecular defects across multiple bone cell populations in a mouse model of CKD, and our collective results suggest a potentially earlier and more broad bone pathology in this disease than previously recognized.

cell biology↗
bioRxiv2022

Structure of the histone acetyltransferase NuA4 complex

NuA4, one of two major histone acetyltransferase complexes in Saccharomyces cerevisiae, specifically acetylates histone H4 and H2A, and therefore loosen the histone-DNA contacts, resulting in increased transcriptional activity. Here we present a near-atomic resolution structure of NuA4 complex through cryonic electron microscopy. The structure comprises six subunits and a total of 5,000 amino acids, within which Eaf1 and Eaf2 twist against each other and span from Actin-Arp4 module to the platform subunit Tra1, forming the backbone of NuA4. The locations of four missing components including Yaf9, and Esa1, Yng2 and Eaf6 of Piccolo module were indicated based on the structure. Intriguingly, our biochemical data showed that NuA4 doesnt bind nucleosome in vitro, whereas possesses strong histone acetyltransferase activity. Its Piccolo module, which has long been regarded as the catalytic module of NuA4, acetylates the nucleosome substrate at almost the same efficiency, but is capable of binding nucleosome. Combining the structural and biochemical data give rise to a model in which the entire NuA4 adopts an auto-inhibited conformation, and undergoes structural rearrangements upon substrate and co-factor additions, restoring its capacity of nucleosome-binding and activity of hyper-acetylation.

biochemistry↗
bioRxiv2022

Underlying beneficial effects of Rhubarb on constipation-induced inflammation, disorder of gut microbiome and metabolism

BackgroundAlthough constipation is a common syndrome and a worldwide health problem. Constipation patients are becoming younger with a 29.6% overall prevalence in the children, which has captured great attention because of its epigenetic rejuvenation and recurrent episodes. Despite the usage of rhubarb to relieve constipation, novel targets and genes involved in target-relevant pathways with remarkable functionalities should still be sought after. Materials and methodsWe established a reliable constipation model in C57B/6N male mice using intragastric administration diphenoxylate and the eligible subjects received 600mg/25g rhubarb extraction to ameliorate constipation. Resultant constipation was morphological and genetically compared with the specimen from different groups. ResultsThe constipation mice exhibited thicker muscle layers, improved content of cytokines, including IL-17 and IL-23, and lower content of IL-22. The bacterial abundance and diversity varied tremendously. Notably, the alterations were reversed after rhubarb treatment. Additionally, SCFA and MLCFA were significantly influenced by constipation accompanied by enhanced expressions of SCFA receptors, GPR41 and GPR43. ConclusionThis thesis has provided an insight that rhubarb promoted the flexibility of collagen fiber, reduced pro-inflammatory cytokines and enhanced anti-inflammatory cytokines, and maintained intestinal microflora balance with potential effects on affecting the metabolism of fatty acids and polyamines.

pharmacology and toxicology↗
bioRxiv2022

Locus c{oelig}ruleus noradrenergic neurons phase-lock to prefrontal cortical and hippocampal infra-slow rhythms which synchronize with behavioral events

The locus cruleus (LC) is the primary source of noradrenergic projections to the forebrain, and, in prefrontal cortex, is implicated in decision-making and executive function. LC neurons phase-lock to cortical infra-slow wave oscillations during sleep. Such infra-slow rhythms are rarely reported in awake states, despite their interest, since they correspond to the time scale of behavior. Thus, we investigated LC neuronal synchrony with infra-slow rhythms in awake rats performing an attentional set-shifting task. Local field potential (LFP) oscillation cycles in prefrontal cortex and hippocampus on the order of 0.4 Hz phase-locked to task events at crucial maze locations. Indeed, successive cycles of the infra-slow rhythms showed different wavelengths, as if they are periodic oscillations that can reset phase relative to salient events. Simultaneously recorded infra-slow rhythms in prefrontal cortex and hippocampus could show different cycle durations as well suggesting independent control. Most LC neurons (including optogenetically identified noradrenergic neurons) recorded here were phase-locked to these infra-slow rhythms, as were hippocampal and prefrontal units recorded on the LFP probes. The infra-slow oscillations also phase-modulated gamma amplitude, linking these rhythms at the time scale of behavior to those coordinating neuronal synchrony. This would provide a potential mechanism where noradrenaline, released by LC neurons in concert with the infra-slow rhythm, would facilitate synchronization or reset of these brain networks, underlying behavioral adaptation.

neuroscience↗
bioRxiv2022

A Unified Catalog of 19,251 Non-human Reference Species Genomes Provides New Insights into the Mammalian Gut Microbiomes

The gut microbiota is essential for host health and survival. Here, using samples from animals living in the Qinghai-Tibetan Plateau, we recovered 119,568 metagenome-assembled genomes (MAGs) that were clustered into 19,251 species-level genome bins (SGBs) of which most represent novel species. We present a novel mechanism shaping mammalian gut microbiomes using ancestral founder bacteria (AFB) as a core skeleton and recurring lineage-specific gains of microbial species that are transferred frequently among multiple hosts, not strictly limited by host phylogeny. Such lineage specific gains are responsible for increasing gut microbial diversity, maintaining functional stability, and endowing specific functions for host adaptions. Our analyses did not support the existence of co-phylogeny or co-speciation events between mammal hosts and their individual gut symbionts. The results presented in this study not only reveal novel unique gut microbial species and offer insight of value for understanding the diversity, stability, functionality of the mammalian gut microbiomes, and the co-evolution with their hosts, but also emphasize that animals living in extreme environments are a promising resource for the discovery of novel biological functions.

microbiology↗
bioRxiv2022

Hepatic AMPK activation in response to dynamic REDOX balance is a biomarker of exercise to improve blood glucose control

Antioxidant intervention is considered to inhibit reactive oxygen species (ROS) and alleviates hyperglycemia. Paradoxically, moderate exercise can produce ROS to improve diabetes. The exact redox mechanism of these two different approaches remains largely unclear. Here, by comparing exercise and antioxidants intervention on type 2 diabetic rats, we found moderate exercise upregulated compensatory antioxidant capability and reached a higher level of redox balance in the liver. In contrast, antioxidant intervention achieved a low-level redox balance by inhibiting oxidative stress. Both of these two interventions could promote glycolysis and aerobic oxidation mediated by hepatic AMPK activation, ameliorating diabetes. During exercise, different levels of ROS generated by exercise have differential regulations on the activity and expression of hepatic AMPK. Moderate exercise-derived ROS promoted hepatic AMPK glutathionylation activation. However, excess exercise increased oxidative damage, and inhibited the activity and expression of AMPK. Overall, our results illustrate that both exercise and antioxidant intervention improve blood glucose in diabetes by promoting redox balance, despite the levels of redox balance are different. Moreover, the activation and expression of AMPK could act as a biomarker to reflect the effective treatment range for diabetes. This finding provides theoretical evidence for the precise regulation of diabetes by antioxidants and exercise.

biochemistry↗
bioRxiv2022

Identification of transcriptional network disruptions in drug-resistant prostate cancer with TraRe

Metastatic castration-resistant prostate cancer (mCRPC) presents very low survival rates due to lack of response or acquired resistance to the available therapies. To date no molecular mechanisms of resistance have been identified, pointing out their complex dynamics. To identify key genes and processes associated with phenotypically-driven regulatory differences, we developed TraRe, a computational method that provides a three-tier analysis: i) at the network level, inferring differentially regulated modules; ii) at the regulon level, identifying regulatory relationships linked to phenotypic differences; and iii) at the single gene level, identifying TFs consistently linked to rewired modules. We applied TraRe (available in Bioconductor with full documentation) to transcriptomic data from 46 mCRPC patients with Abiraterone-response clinical data and uncovered abrogated immune response regulatory modules that showed strong differential regulation in Abi-resistant patients. These modules were replicated in an independent mCRPC study. Further, we experimentally validated key rewiring predictions and their associated transcription factors. Among them, ELK3, MXD1, and MYB were found to have a differential role in cell survival for Abi-response-specific settings. Moreover, we identified the role of ELK3 in cell migration capacity, which could have direct impact on mCRPC. Collectively, these findings shed light on the underlying regulatory mechanisms driving abiraterone response.

bioinformatics↗
bioRxiv2022

Structural and functional characteristics of SARS-CoV-2 Omicron subvariant BA.2 spike

The Omicron subvariant BA.2 has become the dominant circulating strain of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in many countries. We have characterized structural, functional and antigenic properties of the full-length BA.2 spike (S) protein and compared replication of the authentic virus in cell culture and animal model with previously prevalent variants. BA.2 S can fuse membranes more efficiently than Omicron BA.1, mainly due to lack of a BA.1-specific mutation that may retard the receptor engagement, but still less efficiently than other variants. Both BA.1 and BA.2 viruses replicated substantially faster in animal lungs than the early G614 (B.1) strain in the absence of pre-existing immunity, possibly explaining the increased transmissibility despite their functionally compromised spikes. As in BA.1, mutations in the BA.2 S remodel its antigenic surfaces leading to strong resistance to neutralizing antibodies. These results suggest that both immune evasion and replicative advantage may contribute to the heightened transmissibility for the Omicron subvariants.

microbiology↗
bioRxiv2022

Glucocorticoids Mediate Transcriptome-wide Alternative Polyadenylation: Mechanisms and Functional Implications

Alternative polyadenylation (APA) is a common genetic regulatory mechanism that generates distinct 3' ends for RNA transcripts. Changes in APA have been associated with multiple biological processes and disease phenotypes. However, the role of hormones and their drug analogs in APA remains largely unknown. In this study, we investigated transcriptome-wide the impact of glucocorticoids on APA in 30 human B-lymphoblastoid cell lines. We found that glucocorticoids could regulate APA of a subset of genes, possibly by changing the expression of 142 RNA-binding proteins, some with known APA-regulating properties. Interestingly, genes with glucocorticoid-mediated APA were enriched in viral translation-related pathways, while genes with glucocorticoid-mediated expression were enriched in interferon and interleukin pathways, suggesting that glucocorticoid-mediated APA might result in functional consequences distinct from gene expression. Glucocorticoid-mediated APA was also cell-type-specific, suggesting an action of glucocorticoids that may be unique to immune regulation. We also observed evidence for genotype-dependent glucocorticoid-mediated APA, providing potential functional mechanisms for a series of common genetic variants that had previously been associated with immune disorders, but without a clear mechanism. In summary, this study reports a series of novel observations regarding the impact of glucocorticoids on APA, suggesting novel avenues for mechanistic studies of hormonal biology.

genomics↗
bioRxiv2022

Mosaic RBD nanoparticles protect against multiple sarbecovirus challenges in animal models

To combat future SARS-CoV-2 variants and spillovers of SARS-like betacoronaviruses (sarbecoviruses) threatening global health, we designed mosaic nanoparticles presenting randomly-arranged sarbecovirus spike receptor-binding domains (RBDs) to elicit antibodies against conserved/relatively-occluded, rather than variable/immunodominant/exposed, epitopes. We compared immune responses elicited by mosaic-8 (SARS-CoV-2 and seven animal sarbecoviruses) and homotypic (only SARS-CoV-2) RBD-nanoparticles in mice and macaques, observing stronger responses elicited by mosaic-8 to mismatched (not on nanoparticles) strains including SARS-CoV and animal sarbecoviruses. Mosaic-8 immunization showed equivalent neutralization of SARS-CoV-2 variants including Omicron and protected from SARS-CoV-2 and SARS-CoV challenges, whereas homotypic SARS-CoV-2 immunization protected only from SARS-CoV-2 challenge. Epitope mapping demonstrated increased targeting of conserved epitopes after mosaic-8 immunization. Together, these results suggest mosaic-8 RBD-nanoparticles could protect against SARS-CoV-2 variants and future sarbecovirus spillovers.

microbiology↗
bioRxiv2022

TGF-beta1 induced S100 family protein expression is associated with epithelial to mesenchymal transition states and poor survival in pancreatic cancer

Epithelial-mesenchymal transition (EMT) is a continuum that includes epithelial, partial EMT (P-EMT) and mesenchymal states, each of which are associated with cancer progression, invasive capabilities and ultimately metastasis. We have employed a lineage traced sporadic model of pancreatic cancer to generate a murine organoid biobank from primary and secondary tumors, including sublines that have undergone P-EMT and complete EMT (C-EMT). Using an unbiased proteomics approach, we found that the morphology of the organoids predicts the EMT state, with solid organoids associated with a P-EMT signature. We also observed that exogenous TGF{beta}1 induces a solid organoid morphology that is associated with changes in the S100 family, C-EMT and the formation of high-grade tumors. S100A4 may represent a useful biomarker to predict EMT state, disease progression and outcome for pancreatic cancer patients.

cancer biology↗
bioRxiv2022

Comprehensive dissection of immune microenvironment in the progression of early gastric cancer at spatial and single-cell resolution

While the changes of tumor immune microenvironment (TME) have critical implications for most tumor progression, works that could reveal the compositions and immunity features of TME are needed. Profiling gastric malignant cells at single-cells resolution has shown the transcriptional heterogeneity is represented at different states of gastric cancer, implying that diverse cell states may exist, including immune cells, and all components in TME make some balances in early gastric cancer (EGC) progression. However, it remains unclear how immune cells contributing malignancy of gastritis, constituting general characteristics of gastric TME. Furthermore, the role of specific interactions among cells in gastric TME remains to be illustrated. Here, we performed spatial transcriptomes and single-cell RNA-seq analysis across 18 gastric samples, identifying 17 celltypes and reconstructing their location information. We found that immune cells represented different degree of dysregulations during the progression from non-atrophic gastritis (NAG), atrophic gastritis (AG) to EGC, including imbalance of cytotoxic and inhibitory effects in T cells, maturation inhibition in B cells and malignant genes up-regulated obviously in myeloid cells. Besides, pathway activities showed that hypoxia, reactive oxygen species and fatty metabolism signaling were activated from AG stage, which may accelerate progression of EGC. Moreover, cellular interactions further identified the roles of hypoxia in gastric TME. Overall, the multi-omics data presented in this study offer a comprehensive view of immune cell types, states changes and locations within the gastric tissues during the progression from NAG, AG to EGC, advancing our understanding of the composition and immunity of different gastric states, offering diagnostic and preventive thoughts for EGC.

cancer biology↗
bioRxiv2022

Improving the Identification Coverage of Protein Interactome by Enhancing the Click Chemistry-based Cross-linking Enrichment Efficiency

Chemical cross-linking coupled with mass spectrometry has emerged as a powerful strategy which enables global profiling of protein interactome with direct interaction interfaces in complex biological systems. The alkyne-tagged enrichable cross-linkers are preferred to improve the coverage of low-abundance cross-linked peptides, combined with click chemistry for biotin conjugation to allow the cross-linked peptides enrichment. However, a systematic evaluation on the efficiency of click approaches (protein-based or peptide-based) and diverse cleavable click chemistry ligands (acid, reduction, photo) for cross-linked peptides enrichment and release is lacking. Herein, together with in vivo chemical cross-linking by alkyne-tagged cross-linker, we explored the click chemistry-based enrichment approaches on protein and peptide level with three cleavable click chemistry ligands, respectively. By comparison, the approach of protein-based click chemistry conjugation with acid-cleavable tag was demonstrated to permit the most cross-linked peptides identification. The advancement of this strategy enhanced the proteome-wide cross-linking analysis, permitting a detection of 5,017 protein-protein interactions among 1,909 proteins across all subcellular compartments with wide abundance distribution in cell. Therefore, all these results demonstrated a guideline value of our work for efficient cross-linked peptides enrichment, thus facilitated the in-depth profiling of protein interactome for functional analysis. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/475819v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@1a7ae4forg.highwire.dtl.DTLVardef@1a4d6d2org.highwire.dtl.DTLVardef@113a0eborg.highwire.dtl.DTLVardef@1a9c193_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗
bioRxiv2022

Spatially resolved profiling of protein conformation and interactions by biocompatible chemical cross-linking in living cells

The protein structures and interactions that maintain and regulate cellular processes in different subcellular organelles are heterogeneous and dynamic. However, it remains challenging to characterize the subcellular specificity and translocation of protein complexes in terms of conformation and interactions. Herein, we developed a spatially resolved protein complex profiling approach by biocompatible chemical cross-linking in living cells (SPACX) to monitor the dynamics of protein conformation, interactions and translocation. The advancement of fast capturing protein complexes in the physiological state, coupled with efficient enrichment of the cross-linked peptides, ensured deep-coverage analysis of the protein interactome in living cells. By ensemble structure refinement with cross-linking restraints, subcellular-specific conformation heterogeneity was identified for PTEN. PTEN displayed a broader range of dynamic conformation changes on the dual specificity domains in the nucleus than in the cytoplasm. Moreover, based on conformational differences, different interacting assemblies involving 25 cytoplasm-exclusively and 18 nucleus-exclusively PTEN-interacting proteins were found to account for diverse biological functions. Upon ubiquitin-proteasome system (UPS) stress, the assembly of PTEN and its interacting partners changed obviously during translocation. We newly identified 36 PTEN-interacting proteins, which were found to be highly enriched in functional pathways closely related to cell apoptosis. Inspiringly, the interactions among PTEN isoforms and their interacting proteins were accessible by the determination of sequence-unique cross-linking interfaces for direct interactions. All these results indicate the promise of SPACX to elucidate the functional heterogeneity of proteins in individual subcellular sociology.

biochemistry↗
bioRxiv2022

Cyanopeptides Restriction and Degradation Co-mediate Microbiota Assembly During a Freshwater Cyanobacterial Harmful Algal Bloom (CyanoHAB)

Cyanobacterial harmful algal blooms (CyanoHABs) are globally intensifying and exacerbated by climate change and eutrophication. However, microbiota assembly mechanisms underlying CyanoHABs remain scenario specific and elusive. Especially, cyanopeptides, as a group of bioactive secondary metabolites of cyanobacteria, could affect microbiota assembly and ecosystem function. Here, the trajectory of cyanopeptides were followed and linked to microbiota during Microcystis-dominated CyanoHABs in lake Taihu, China. The most abundant cyanopeptide classes detected included microginin, spumigin, microcystin, nodularin and cyanopeptolin with total MC-LR-equivalent concentrations between 0.23 and 2051.54 ppb, of which cyanotoxins beyond microcystins (e.g., cyanostatin B and nodularin_R etc.) far exceeded reported organismal IC50 and negatively correlated with microbiota diversity, exerting potential collective eco-toxicities stronger than microcystins alone. The microbial communities were differentiated by size fraction and sampling date throughout CyanoHABs, and surprisingly, their variances were better explained by cyanopeptides (19-38%) than nutrients (0-16%). Cyanopeptides restriction (e.g., inhibition) and degradation are first quantitatively verified as the deterministic drivers governing community assembly, with stochastic processes being mediated by interplay between cyanopeptide dynamics and lake microbiota. This study presents an emerging paradigm in which cyanopeptides restriction and degradation co-mediate lake water microbiota assembly, unveiling new insights about the ecotoxicological significance of CyanoHABs to freshwater ecosystems.

ecology↗