Biology subjects

Zhou, H.

Publications and source records attributed to Zhou, H..

At least 271 records · Page 15Linked to original sources

bioRxiv2023

Landscape of IGH germline genes of Chiroptera and the pattern of Rhinolophus affinisbat IGH CDR3 repertoire

The emergence and re-emergence of a number of viruses from bats that impact human and animal health has resulted in a resurgence of interest in bat immunology. Characterizing the immune receptor repertoire is critical to understanding how bats coexist with viruses in the absence of disease and developing new therapeutics to target viruses in humans and susceptible livestock. We annotated IGH germline genes of Rhinolophus ferrumequinum (RF), Phyllostomus discolor (RD) and Pipistrellus pipistrellus (PP), and investigated the evolutionary relationship between bat germline genes and that of human, mouse, cow, and dog. The IGH repertoire characteristics of Rhinolophus affinis bat (RA) were also analyzed. The V gene families of all three bat species can be classified into three Clan, although PD is special with the abnormal length of IGH locus and 22 reverse V genes. Moreover, the bats germline genes are quite differed from those of human, mouse, cow, and dog in evolution, but the three bat species have high homology. The CDR3 repertoire of RA are unique in many aspects including CDR3 subclass, V/J genes access and pairing, CDR3 clones and somatic high-frequency mutation compared with that of human and mouse, which may be the immunologic basis for the asymptomatic nature of viral infection in bats. This study provide immune genome information and extensive reference for the basic research of bat and virus infection mechanism.

immunology↗

bioRxiv2023

Learning functional conservation between pig and human to decipher evolutionary mechanisms underlying gene expression and complex trait

The assessment of genomic conservation between human and pig at the functional level can help understand and improve the potential of pig as a human biomedical model. To address this, we developed a Deep learning-based approach to learn the Genomic Conservation at the Functional level (DeepGCF) between species by integrating 386 and 374 epigenome and transcriptome profiles from human and pig, respectively. DeepGCF demonstrated a better prediction performance compared to the previous functional conservation prediction method. In addition, we showed that the resulting DeepGCF score captures the functional conservation by examining DeepGCF on chromatin states, sequence ontologies, and regulatory variants. Regions with higher DeepGCF score play a more important role in regulatory activities and show heritability enrichment in human complex traits and diseases. Our DeepGCF approach shows a promising application on the comparison of cross-species functional conservation, and the model framework can be easily adapted to other species. By expanding the model to integrate the functional profiles of multiple species, including human, mouse, pig, cattle, and other livestock animals in the future, the functional conservation information will provide additional insight into the genetic and evolutionary mechanisms behind complex traits and diseases.

genetics↗

bioRxiv2023

3D co-culture of macrophages and fibroblasts in a sessile drop chip for unveiling the role of macrophages in skin wound-healing

Three-dimension (3D) cell co-cultural spheroids exhibit enhanced cellular functions and they can mirror in-vivo microenvironments. Herein, a sessile drop chip was developed to construct 3D spheroids for mirroring the wound healing microenvironment. The sessile drop chip holds the superhydrophobic surface of each microwell which can facilitate cell suspensions transfer to spheroids through the offset of surface tension and gravity, and each microwell has a cylinder hole that offers adequate oxygen to spheroids. It was demonstrated that the 3T3 fibroblast spheroid and the 3T3 fibroblast/M2-type macrophage co-culture spheroid can be formed and remained the physiological activity within nine days. 3D morphology of spheroids was reconstructed using the transparent processing technology and Z-stack function of confocal microscopy. Characteristics of proliferation and differentiation were analyzed by using nano antibody-based 3D immunostaining assay. Results revealed that M2-type macrophages can promote the proliferation and differentiation of the 3T3 fibroblast spheroid. This study presented a novel affordable platform for developing 3D spheroids and provides a 3D model for investigating the macrophages-associated wound healing microenvironment.

bioengineering↗

bioRxiv2023

ACE2 Receptor Decoy is a Potent Prophylactic and Therapeutic for SARS-CoV-2

The emergence of SARS-CoV-2 variants with highly mutated spike proteins has presented an obstacle to the use of monoclonal antibodies for the prevention and treatment of SARS-CoV-2 infection. We show that a high affinity receptor decoy protein in which a modified ACE2 ectodomain is fused to a single domain of an immunoglobulin heavy chain Fc region dramatically suppressed virus loads in mice upon challenge with a high dose of parental SARS-CoV-2 or Omicron variants. The decoy also potently suppressed virus replication when administered shortly post-infection. The decoy approach offers protection against the current viral variants and, potentially, against SARS-CoV-2 variants that may emerge with the continued evolution of the spike protein or novel viruses that use ACE2 for virus entry.

microbiology↗

bioRxiv2022

The ribosome-associated chaperone Zuo1 controls translation upon TORC1 inhibition

Eukaryotic cells achieve proteostasis by ensuring their protein requirements are met through tight control of TORC1 activity. Upon TORC1 inhibition, degradative activity is increased, and protein synthesis is reduced through inhibition of translation initiation, to maintain cell viability. Here, we show that the ribosome-associated complex (RAC)/Ssb chaperone system is required to maintain proteostasis and cell viability under TORC1 inhibition, in yeast. In the absence of the Hsp40 cochaperone Zuo1, translation does not decrease in response to loss of TORC1 activity. The functional interaction between Zuo1 and its Hsp70 partner, Ssb, is required for proper translational control and proteostasis maintenance upon TORC1 inhibition. Further, we have found that the rapid degradation of eIF4G following TORC1 inhibition is prevented in zuo1{Delta} cells, contributing to decreased survival in these conditions. Our findings suggest a new role for RAC/Ssb in regulating translation in response to changes in TORC1 signalling.

molecular biology↗

bioRxiv2022

New insight into the Germline Genes and CDR3 Repertoire of the TCRβ chain in Chiroptera

Bats are known to be natural reservoirs of many viruses, yet their unique immune system enables them to coexist with viruses without frequently exhibiting disease symptoms. The current understanding of the bat adaptive immune system is limited, as there is no database or tool capable of processing T-cell receptor (TCR) sequences for bats, and the diverse nature of the Chiroptera. We established a reference library of TCR-{beta} germline genes by annotating three Chiroptera: The Greater Horseshoe Bat (Rhinolophus ferrumequinum, R. ferrumequinum), The Pale spear-nosed Bat (Phyllostomus discolor, P. discolor), and the Common Pipistrelle (Pipistrellus pipistrellus, P. pipistrellus). The distinct variations in the distribution of TRBV genes among the three types of bats could have a direct impact on the diversity of the TCR repertoire, as evidenced by the presence of conserved amino acids that indicate the T-cell recognition of antigens in bats is MHC-restricted. Furthermore, we conducted an analysis of the TCR-{beta} repertoire in the Intermediate Horseshoe Bat (Rhinolophus affinis, R. affinis) using high-throughput sequencing (HTS). The bats TCR-{beta} repertoire is formed through rearrangement of the V-D-J-C genes, with D-J/V-D deletions and insertion resulting a high diversity.

immunology↗

bioRxiv2022

Generate a new crucian carp (Carassius auratus) strain without intermuscular bones by knocking out bmp6

Elimination of intermuscular bones (IMBs) is vital to the aquaculture industry of cyprinids. In our previous study, we characterized bmp6 as essential in the development of IMBs in zebrafish. Knockout of bmp6 results in the absence of IMBs in zebrafish without affecting growth and reproduction. Therefore, we hypothesized that bmp6 could be used to generate new cyprinid strains without IMBs by gene editing. In this study, we established a gene editing strategy for knocking out the two orthologs of bmp6 in diploid crucian carp (Carassius auratus). We obtained an F3 population with both orthologs knocked out, in which no IMBs were detected by bone staining and X-ray, indicating that the new strain without IMBs (named WUCI strain) was successfully generated. Furthermore, we extensively evaluated the performance of the new strain in growth, reproduction, nutrient components, muscle texture and structure, and metabolites in muscle. The results showed that the WUCI strain grew faster than the wild-type crucian carp at 4-month-age. The reproductive performance and flesh quality did not show significant differences between the WUCI strain and wild-type crucian carp. Moreover, the metabolomics analysis suggested that the muscle tissues of the WUCI strain significantly enriched some metabolites belonging to the Thiamine metabolism, Nicotinate and Nicotinamide Metabolism pathway, which plays beneficial effects in anti-aging, anti-oxidant, and anti-radiation damage. In conclusion, we established a strategy to eliminate IMBs in crucian carp and obtain a WUCI strain whose performance was expected compared to the wild-type crucian carp; meanwhile, the WUCI strain enriched some beneficial metabolites to human health in muscle tissue. This study is the first report that a farmed cyprinid strain without IMBs, which could be stably inherited, was obtained worldwide; it provided excellent germplasm for the cyprinids aquaculture industry and a useful molecular tool for eliminating IMBs in other cyprinids.

genetics↗

bioRxiv2022

The Cul5 E3 Ligase Complex Is a Key Negative Feedback Regulator of TCR/IL2 Signaling and Anti-Tumor Activity in CD8+ T Cells

CD8+ T cells play an important role in tumor immune surveillance and control. Better understanding of the regulation of their anti-tumor actions and improving their cytotoxic function and persistence will help advancing cancer immunotherapies. Here, we report the development of a step-wise CRISPR knockout (KO) screening strategy under the selection of TGF-{beta}, a clinically relevant immunosuppressive pressure. The screen identifies Cul5 as a negative-feedback regulator of the core signaling pathways, differentiation, and persistence of CD8+ T cell. Cul5 KO in mouse CD8+ T cells significantly improves their tumor control ability in vitro and in vivo with significant proteomic alterations that generally enhance TCR and cytokine signaling, effector function, stemness, and survival of CD8+ T cell. Mechanistically, Cul5, whose protein content and active, neddylated form increase upon TCR-stimulation, interacts with SOCS-box-containing Pcmtd2 and negatively regulates TCR and IL2/STAT5 signaling by decreasing TCR and IL2 signaling molecules. Moreover, Cul5 KO in human CD8+ T cells phenocopies that in mouse CD8+ T cells. Furthermore, KO of CTLA4 that is markedly upregulated by Cul5 KO in mouse and human CD8+ cells further enhances anti-tumor effect of Cul5 KO, and a neddylation inhibitor enhances CD8 effector activities largely dependently of Cul5. These results together not only reveal a previously unknown negative-feedback regulatory mechanism for CD8+ T cells, but also have strong translational implications in cancer immunotherapy.

immunology↗

bioRxiv2022

Inflammation in the tumor-adjacent lung as a predictor of clinical outcome in lung adenocarcinoma

Early-stage lung adenocarcinoma is typically treated by surgical resection of the tumor. While in the majority of cases surgery can lead to cure, approximately 30% of patients progress. Despite intense efforts to map the genetic landscape of early-stage lung tumors, there has been limited success in discovering accurate biomarkers that can predict clinical outcomes. Meanwhile, the role of the tumor-adjacent tissue in cancer progression has been largely ignored. To test whether tumor-adjacent tissue can be informative of progression-free survival and to probe the underlying molecular pathways involved, we designed a multi-omic study in both tumor and matched tumor-adjacent histologically normal lung tissue from the same patient. Our study includes 143 treatment naive stage I cases with long-term patient follow-up and is, to our knowledge, the largest such study with the longest follow-up. We performed a comprehensive histologic characterization of all tumors, mapped the mutational landscape and probed the transcriptome of both tumor and adjacent normal tissue. We evaluated the predictive power of each data modality and showed that the transcriptome of tumor-adjacent histologically normal lung tissue is the only reliable predictor of clinical outcome. Unbiased discovery of co-expressed gene modules revealed that inflammatory pathways are upregulated in the tumor-adjacent tissue of patients at high risk for disease progression. Furthermore, single-cell transcriptome analysis in the tumor-adjacent lung demonstrated that progression-associated inflammatory signatures were broadly expressed by both immune and non-immune cells including mesothelial cells, alveolar type 2 cells and fibroblasts, CD1 dendritic cells and MAST cells. Collectively, our studies suggest that molecular profiling of tumor-adjacent tissue can identify patients that are at high risk for disease progression.

bioinformatics↗

bioRxiv2022

A novel antidepressant acting via allosteric inhibition of GluN2D-incorporated NMDA receptors at GABAergic interneurons

N-methyl-D-aspartate receptors (NMDARs) are glutamate-gated calcium-permeable excitatory channels. They have attracted great interest as potential targets for the treatment of depression in recent years. NMDARs typically assemble as heterotetramers composed of two GluN1 and two alternative GluN2 (2A-2D) subunits, the latter of which endow various subtypes with diverse gating and pharmacological properties. To date, limited molecules with GluN2 specificity have been identified to show antidepressant effects. Here, we identify a compound termed YY-23 extracted from Rhizoma Anemarrhenae allosterically inhibited the channel activities of GluN2C- or GluN2D-incorporated NMDARs, an effect that was presumably influenced by the S2 segment in the ligand-binding domain of the GluN2 subunit. We found that prefrontal GluN2D-containing NMDARs were predominantly expressed at GABAergic interneurons rather than pyramidal neurons. Furthermore, we revealed that YY-23 suppressed the activity of GluN2D-containing NMDARs and GABAergic neurotransmission in the medial prefrontal cortex (mPFC). As a consequence, this GABAergic disinhibition facilitated the excitatory transmission. Behavioural experiments showed that YY-23 acted as a rapid antidepressant in both stress-naive and stressed animal models, which was abolished in Grin2d-knockout mice. Together, our findings suggest that GluN2D-incorporated NMDARs on GABAergic interneurons might be promising therapeutic targets for the treatment of depression.

neuroscience↗

bioRxiv2022

Breast cancer progression and metastasis to lymph nodes reveals cancer cell plasticity and MHC class II-mediated immune regulation

Tumor-draining lymph nodes are critical sites for generating tumor antigen-specific T cells and are associated with durable immune responses. However, lymph nodes are often the first site of metastasis and lymph node metastases portend worse outcomes. Through cross-species single cell gene expression analysis of breast cancer progression and metastasis to lymph nodes, we uncovered features that define the heterogeneity, plasticity, and immune evasion of cancer cells. Notably, a subpopulation of metastatic cancer cells in the lymph node were marked by high levels of MHC class II (MHC-II) gene expression both in mice and humans. Mechanistically, the IFN-{gamma} and JAK/STAT signaling pathways mediate MHC-II expression in cancer cells. Ablation of IFNGR1/2 or CIITA, the transactivator of MHC-II, in cancer cells prevented tumor progression. Interestingly, MHC-II+ cancer cells lacked co-stimulatory molecule expression, engendered the expansion of regulatory T cells and blunted CD4+ effector T cells in the tumor-draining lymph nodes and favor tumor progression. Overall, our data suggests that cancer cell plasticity during breast cancer progression and metastasis to lymph nodes endows metastatic cells with the ability to avoid immune surveillance. These data provide the basis for new opportunities to therapeutically stimulate anti-cancer immune responses against local and systemic metastases.

cancer biology↗

bioRxiv2022

CRISPR/FnCas12a-mediated efficient multiplex and iterative genome editing in bacterial plant pathogens without donor DNA templates

CRISPR-based genome editing technology is revolutionizing prokaryotic research, but it has been rarely studied in bacterial plant pathogens. Here, we have developed a targeted genome editing method with no requirement of donor templates for convenient and efficient gene knockout in Xanthomonas oryzae pv. oryzae (Xoo), one of the most important bacterial pathogens on rice, by employing the heterogenous CRISPR/Cas12a from Francisella novicida and NHEJ proteins from Mycobacterium tuberculosis. FnCas12a nuclease generated both small and large DNA deletions at the target sites as well as it enabled multiplex genome editing, gene cluster deletion and plasmid cure in the Xoo PXO99A strain. Accordingly, a non-TAL effector-free polymutant strain PXO99AD25E, which lacks all 25 Xop genes involved in Xoo pathogenesis, has been engineered through iterative genome editing. Whole-genome sequencing analysis indicated that FnCas12a did not have a noticeable off-target effect. In addition, we revealed that these strategies are also suitable for targeted genome editing in another bacterial plant pathogen Pseudomonas syringae pv. tomato (Pst). We believe that our bacterial genome editing method will greatly expand the CRISPR study on microorganisms and advance our understanding of the physiology and pathogenesis of Xoo.

microbiology↗

bioRxiv2022

Age-specific activation of microglial genes in a mouse model of AD

Alzheimers disease (AD) is a neurodegenerative disease that increases its prevalence with age. AD is characterized by a progressive loss of cognitive function, leading to dementia. The conventional pathological hallmarks of AD include deposition of amyloid plaque, aggregation of neurofibrillary tangles, and sustained inflammation. Recent studies have confirmed that immune responses play an important role in AD progression. Single-cell RNA-sequencing (RNA-seq) studies of the AD mouse model identified the disease-associated microglia (DAM) population during AD progression, where the pro-inflammatory activity was mediated by Trem2. However, the time points and nature of DAM activation remain largely unknown. Here, we compared the time-result bulk sequencing andsingle nuclei RNA sequencing data in a mouse model of AD. We identified an age-dependent activation of Trem2 targeted immune-related genes. We showed that Trem2 affected DAM activation at an early stage (4-8 months) rather than the late stage (12-16 months). Our results highlight an age-dependent change in the Trem2-linked immune response in AD.

neuroscience↗

bioRxiv2022

Unsupervised Discovery of Ancestry Informative Markers and Genetic Admixture Proportions in Biobank-Scale Data Sets

Admixture estimation plays a crucial role in ancestry inference and genomewide association studies (GWAS). Computer programs such as ADMIXTURE and STRUCTURE are commonly employed to estimate the admixture proportions of sample individuals. However, these programs can be overwhelmed by the computational burdens imposed by the 105 to 106 samples and millions of markers commonly found in modern biobanks. An attractive strategy is to run these programs on a set of ancestry informative SNP markers (AIMs) that exhibit substantially different frequencies across populations. Unfortunately, existing methods for identifying AIMs require knowing ancestry labels for a subset of the sample. This supervised learning approach creates a chicken and the egg scenario. In this paper, we present an unsupervised, scalable framework that seamlessly carries out AIM selection and likelihood-based estimation of admixture proportions. Our simulated and real data examples show that this approach is scalable to modern biobank data sets. Our implementation of the method is called OpenADMIXTURE.

bioinformatics↗

bioRxiv2022

Multi-trait genome-wide association analyses leveraging alcohol use disorder findings identify novel loci for smoking behaviors in the Million Veteran Program

Smoking behaviors and alcohol use disorder (AUD), moderately heritable traits, commonly co-occur in the general population. Single-trait genome-wide association studies (GWAS) have identified multiple loci for smoking and AUD. However, GWASs that have aimed to identify loci contributing to comorbid smoking and AUD have used small samples and thus have not been highly informative. Applying multi-trait analysis of GWASs (MTAG), we conducted a joint GWAS of smoking and AUD with data from the Million Veteran Program (N=318,694). By leveraging GWAS summary statistics for AUD, MTAG identified 21 genome-wide significant (GWS) loci associated with smoking initiation and 18 loci associated with smoking cessation compared to 16 and 8 loci, respectively, identified by single-trait GWAS. The novel loci for smoking behaviors identified by MTAG included those previously associated with psychiatric or substance use traits. Colocalization analysis identified 10 loci shared by AUD and smoking status traits, all of which achieved GWS in MTAG, including variants on SIX3, NCAM1, and near DRD2. Functional annotation of the MTAG variants highlighted biologically important regions on ZBTB20, DRD2, PPP6C, and GCKR that contribute to smoking behaviors. In contrast, MTAG of smoking behaviors and alcohol consumption (AC) did not enhance discovery compared with single-trait GWAS for smoking behaviors. We conclude that using MTAG to augment the power of GWAS enables the identification of novel genetic variants for commonly comorbid phenotypes, providing new insights into their pleiotropic effects on smoking behavior and AUD.

genetics↗

bioRxiv2022

Functional genomics of cattle through integration of multi-omics data

Functional annotation of the bovine genome was performed by characterizing the spectrum of RNA transcription using a multi-omics approach, combining long- and short-read transcript sequencing and orthogonal data to identify promoters and enhancers and to determine boundaries of open chromatin. A total number of 171,985 unique transcripts (50% protein-coding) representing 35,150 unique genes (64% protein-coding) were identified across tissues. Among them, 159,033 transcripts (92% of the total) were structurally validated by independent datasets such as PacBio Iso-seq, ONT-seq, de novo assembled transcripts from RNA-seq, or Ensembl and NCBI gene sets. In addition, all transcripts were supported by extensive independent data from different technologies such as WTTS-seq, RAMPAGE, ChIP-seq, and ATAC-seq. A large proportion of identified transcripts (69%) were novel, of which 87% were produced by known genes and 13% by novel genes. A median of two 5 untranslated regions was detected per gene, an increase from Ensembl and NCBI annotations (single). Around 50% of protein-coding genes in each tissue were bifunctional and transcribed both coding and noncoding isoforms. Furthermore, we identified 3,744 genes that functioned as non-coding genes in fetal tissues, but as protein coding genes in adult tissues. Our new bovine genome annotation extended more than 11,000 known gene borders compared to Ensembl or NCBI annotations. The resulting bovine transcriptome was integrated with publicly available QTL data to study tissue-tissue interconnection involved in different traits and construct the first bovine trait similarity network. These validated results show significant improvement over current bovine genome annotations.

bioinformatics↗

bioRxiv2022

CryoFIB milling large tissue samples for cryo-electron tomography

Cryo-electron tomography (cryoET), a powerful tool for exploring the molecular structure of large organisms. However, technical challenges still limit cryoET applications on large samples. In particular, locating and cutting out objects of interest from a large tissue sample is an important but difficult step. In this study, we report a sample thinning strategy and workflow for tissue samples based on cryo-focused ion beam (cryoFIB) milling. This workflow provides a full solution for isolating objects of interest by starting from a millimeter-sized tissue sample and ending with hundred-nanometer thin lamellae. The workflow involves sample fixation, pre-sectioning, a two-step milling strategy, and locating the object of interest using cellular secondary electron imaging (CSEI). The two-step milling strategy introduces a coarse milling method to solve the milling efficiency problem for samples as thick as tens of microns, followed by a fine milling method to create a furrow-ridge structure. The furrow-ridge structure guarantees the generation of large, thin lamellae with enhanced mechanical stability and charge-reducing design. CSEI is highlighted in the workflow, which provides conventional, on-the-fly locating during cryoFIB milling. Tests of the complete workflow were conducted to demonstrate the high efficiency and high feasibility of the proposed method.

biophysics↗

bioRxiv2022

Spatiotemporally resolved transcriptomics reveals subcellular RNA kinetic landscape

Spatiotemporal regulation of the cellular transcriptome is crucial for proper protein expression and cellular function1. However, the intricate subcellular dynamics of RNA synthesis, decay, export, and translocation remain obscured due to the limitations of existing transcriptomics methods2-8. Here, we report a spatiotemporally resolved RNA mapping method (TEMPOmap) to uncover subcellular RNA profiles across time and space at the single-cell level in heterogeneous cell populations. TEMPOmap integrates pulse-chase metabolic labeling of the transcriptome with highly multiplexed three-dimensional (3D) in situ sequencing to simultaneously profile the age and location of individual RNA molecules. Using TEMPOmap, we constructed the subcellular RNA kinetic landscape of 991 genes in human HeLa cells from upstream transcription to downstream subcellular translocation. Clustering analysis of critical RNA kinetic parameters across single cells revealed kinetic gene clusters whose expression patterns were shaped by multi-step kinetic sculpting. Importantly, these kinetic gene clusters are functionally segregated, suggesting that subcellular RNA kinetics are differentially regulated to serve molecular and cellular functions in cell-cycle dependent manner. Together, these single-cell spatiotemporally resolved transcriptomics measurements provide us the gateway to uncover new gene regulation principles and understand how kinetic strategies enable precise RNA expression in time and space.

genomics↗