Biology subjects

Yang, H.

Publications and source records attributed to Yang, H..

At least 451 records · Page 25Linked to original sources

bioRxiv2022

HIV-2 Immature Particle Morphology Provides Insights into Gag Lattice Stability and Virus Maturation

Retrovirus immature particle morphology consists of a membrane enclosed, pleomorphic, spherical and incomplete lattice of Gag hexamers. Previously, we demonstrated that human immunodeficiency virus type 2 (HIV-2) immature particles possess a distinct and extensive Gag lattice morphology. To better understand the nature of the continuously curved hexagonal Gag lattice, we have used single particle cryo-electron microscopy with a retrovirus to determine the HIV-2 Gag lattice structure for immature virions. The reconstruction map at 5.5 [A] resolution revealed a stable, wineglass-shaped Gag hexamer structure with structural features consistent with other lentiviral immature Gag structures. Cryo-electron tomography provided evidence for nearly complete ordered Gag lattice structures in HIV-2 immature particles. We also solved a 1.98 [A] resolution crystal structure of the carboxyl-terminal domain (CTD) of the HIV-2 capsid (CA) protein that identified a structured helix 12 supported via an interaction of helix 10 in the absence of the SP1 region of Gag. Residues at the helix 10-12 interface proved critical in maintaining HIV-2 particle release and infectivity. Taken together, our findings provide the first 3D organization of HIV-2 immature Gag lattice and important insights into both HIV Gag lattice stabilization and virus maturation.

microbiology↗

bioRxiv2022

XPD protects CTCF-Cohesin binding sites from somatic mutagenesis

Xeroderma pigmentosum group D (XPD) is a DNA helicase with critical functions in transcription initiation and nucleotide excision repair. Missense mutations in XPD are putative drivers in around 10% of bladder cancers (BLCA), but the associated mutational process remains poorly understood. Here, we examine the somatic mutational landscape of XPD wild-type (n=343) and mutant (n=39) BLCA whole genomes. The genome-wide distribution of somatic mutations is significantly altered in XPD mutants, affecting both APOBEC and non-APOBEC associated mutational processes. Specifically, XPD mutants are enriched in T[C>T]N mutations (SBS2) with altered correlation with replication timing. At a locoregional genomic level, mutant XPD BLCA had striking T>G mutation hotspots at CTCF-cohesin binding sites (CBS) with evidence linking XPD to genomic uracil repair. Leveraging differential distribution of somatic mutations, we developed a machine-learning model for predicting pathogenic XPD mutations, which we validated in an independent TCGA cohort with 100% accuracy. Our model enabled the discovery of missed XPD mutation calls and uncovered pathogenic non-hotspot XPD mutations in bladder cancer. Our study reveals how XPD mutations redistribute somatic mutations in cancer genomes and provides a genome sequencing approach to differentiate driver and passenger XPD mutations.

molecular biology↗

bioRxiv2022

Spatially resolved single-cell deconvolution of bulk RNA-seq using Bulk2Space

Uncovering the tissue molecular architecture at single-cell resolution could help better understand organisms biological and pathological processes. However, bulk RNA-seq can only measure gene expression in cell mixtures, without revealing the transcriptional heterogeneity and spatial patterns of single cells. Herein, we introduce Bulk2Space (https://github.com/ZJUFanLab/bulk2space), a deep learning framework-based spatial deconvolution algorithm that can simultaneously disclose the spatial and cellular heterogeneity of bulk RNA-seq data using existing single-cell and spatial transcriptomics references. The use of bulk transcriptomics to validate Bulk2Space unveils, in particular, the spatial variance of immune cells in different tumor regions, the molecular and spatial heterogeneity of tissues during inflammation-induced tumorigenesis, and spatial patterns of novel genes in different cell types. Moreover, Bulk2Space is utilized to perform spatial deconvolution analysis on bulk transcriptome data from two different mouse brain regions derived from our in-house developed sequencing approach termed Spatial-seq. We have not only reconstructed the hierarchical structure of the mouse isocortex but also further annotated cell types that were not identified by original methods in the mouse hypothalamus.

bioinformatics↗

bioRxiv2022

Structural insights as to how iron-loaded siderophores are imported into Mycobacterium tuberculosis by IrtAB

The ATP-binding cassette (ABC) transporter, IrtAB, plays a vital role in the replication and viability of Mycobacterium tuberculosis (Mtb), where its function is to import iron-loaded siderophores. Unusually, it adopts the typical fold of canonical ABC exporters. Here, we report the structure of unliganded Mtb IrtAB and its structure in complex with ATP, ADP, and an ATP analogue (AMP-PNP) at resolutions from 2.9 to 3.5 [A]. In the ATP-bound state, the two nucleotide-binding domains (NBDs) form a "head-to-tail" dimer, but IrtAB has an unexpectedly occluded conformation, with the inner core forming a large hydrophilic cavity of about 4600 [A]3. Comparison of the structure of the transporter in inward-facing and occluded conformations reveals that the NBD and the intracellular helical region of transmembrane domain (TMD) have an asymmetric allosteric mechanism when ATP binding/hydrolysis such that the one exhibits rigid-body rotation and the other moves in a concerted response as a rigid body. This study provides a molecular basis for the ATP-driven conformational changes that occur in IrtAB and an explanation as to how iron-loaded siderophores are imported into Mtb by IrtAB.

molecular biology↗

bioRxiv2022

Integrating Cell Morphology with Gene Expression and Chemical Structure to Aid Mitochondrial Toxicity Detection

Mitochondrial toxicity is an important safety endpoint in drug discovery. Models based solely on chemical structure for predicting mitochondrial toxicity are currently limited in accuracy and applicability domain to the chemical space of the training compounds. In this work, we aimed to utilize both -omics and chemical data to push beyond the state-of-the-art. We combined Cell Painting and Gene Expression data with chemical structural information from Morgan fingerprints for 382 chemical perturbants tested in the Tox21 mitochondrial membrane depolarization assay. We observed that mitochondrial toxicants differ from non-toxic compounds in morphological space and identified compound clusters having similar mechanisms of mitochondrial toxicity, thereby indicating that morphological space provides biological insights related to mechanisms of action of this endpoint. We further showed that models combining Cell Painting, Gene Expression features and Morgan fingerprints improved model performance on an external test set of 244 compounds by 60% (in terms of F1 score) and improved extrapolation to new chemical space. The performance of our combined models was comparable with dedicated in vitro assays for mitochondrial toxicity. Our results suggest that combining chemical descriptors with biological readouts enhances the detection of mitochondrial toxicants, with practical implications in drug discovery. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=114 SRC="FIGDIR/small/475326v2_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@f29341org.highwire.dtl.DTLVardef@1a1970borg.highwire.dtl.DTLVardef@1c3581dorg.highwire.dtl.DTLVardef@1ce00d4_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO C_FIG

bioinformatics↗

bioRxiv2022

Individual coactivation patterns improve the subject identification and their behavior association

As a complex dynamic system, the brain exhibits spatially organized recurring patterns of activity over time. Coactivation patterns (CAPs), which analyzes data from each single frame, has been utilized to detect transient brain activity states recently. However, previous CAP analyses have been conducted at the group-level, which might neglect meaningful individual differences. Here, we estimate individual CAP states at both subject- and scan-level based on a densely-sampled dataset: Midnight Scan Club. We used differential identifiability, which measures the gap between intra- and intersubject similarity, to evaluate individual differences. We found individual CAPs at the subject-level achieved the best discrimination ability by maintaining high intra-subject similarity and enlarging inter-subject differences, and brain regions of association networks mainly contributed to the identifiability. On the other hand, scan-level CAP states were unstable across scans for the same participant. Expectedly, we found subject-specific CAPs became more reliable and discriminative with more data (i.e., longer duration). As the acquisition time of each participant is limited in practice, our results recommend a data collection strategy that collects more scans with appropriate duration (e.g., 12~15 mins/scan) to obtain more reliable subject-specific CAPs, when total acquisition time is fixed (e.g., 150 mins). Overall, this work has constructed reliable subject-specific CAP states with meaningful individual differences and provides a starting point for the subsequent applications of individual brain dynamics.

neuroscience↗

bioRxiv2022

Phosphoantigens are Molecular Glues that Promote Butyrophilin 3A1/2A1 Association Leading to Vγ9Vδ2 T Cell Activation

Tumor cells and pathogen-infected cells are presented to human {gamma}{delta} T cells based on "inside-out" signaling in which metabolites called phosphoantigens (pAgs) inside target cells are recognized by the intracellular domain of a butyrophilin protein (BTN3A1), leading to an extracellular conformational change. Here, we report that pAgs function as molecular "glues" that initiate a heteromeric association between the intracellular domains of BTN3A1 and the structurally similar BTN2A1. Working with both exogenous and endogenous pAgs, we used x-ray crystallography, mutational studies, cellular assays, synthetic probe as well as molecular dynamics investigations to determine how pAgs glue intracellular BTN3A1 and BTN2A1 together for the "inside-out" signaling that triggers {gamma}{delta} T cell activation. This {gamma}{delta} T cell-specific mode of antigen sensing creates opportunities for the development of alternative immunotherapies against cancer and infectious diseases that do not involve {beta} T cells. One Sentence SummaryThe responses of gamma-delta T cells to cancer cells or pathogens are initiated via the intracellular association of heteromeric butyrophilins that are glued together by isoprenoid metabolites.

immunology↗

bioRxiv2021

High-fidelity Cas13 variants for targeted RNA degradation with minimal collateral effect

CRISPR-Cas13 systems have recently been employed for targeted RNA degradation in various organisms. However, collateral degradation of bystander RNAs has imposed a major barrier for their in vivo applications. We designed a dual-fluorescent reporter system for detecting collateral effects and screening Cas13 variants in mammalian cells. Among over 200 engineered variants, several Cas13 variants (including Cas13d and Cas13X) exhibit efficient on-target activity but markedly reduced collateral activity. Furthermore, transcriptome-wide off-targets and cell growth arrest induced by Cas13 are absent for these variants. Importantly, high-fidelity Cas13 variants show comparable RNA knockdown activity with wild-type Cas13 but no detectable collateral damage in transgenic mice and adeno-associated virus-mediated somatic cell targeting. Thus, high-fidelity Cas13 variants with minimal collateral effect are now available for targeted degradation of RNAs in basic research and therapeutic applications.

molecular biology↗

bioRxiv2021

Transcriptomic Profiling Reveals Cancer-Associated Fibroblasts as Potential Targets for the Prognosis and Treatment of Cervical Squamous Cell Carcinoma

Effective treatment of advanced invasive cervical cancer remains challenging nowadays. Herein, single-nucleus RNA sequencing (snRNA-seq) and SpaTial Enhanced REsolution Omics-sequencing (Stereo-seq) technology are used to investigate the immunological microenvironment of cervical squamous cell carcinoma (CSCC), a major type of cervical cancers. The expression levels of most immune checkpoint genes in tumor and inflammation areas of CSCC were not significantly higher than those in the non-cancer samples except for LGALS9 and IDO1. Stronger signals of CD56+ NK cells and immature dendritic cells are found in the hypermetabolic tumor areas, while more eosinophils, immature B cells, and Treg cells are found in the hypometabolic tumor areas. Moreover, a cluster of cancer-associated fibroblasts (CAFs) are identified around some tumors, which highly expressed ACTA2, POSTN, ITGB4, and FAP. The CAFs might support the growth and metastasis of tumors by inhibiting lymphocyte infiltration and remodeling the tumor extracellular matrix. Furthermore, CAFs are associated with poorer survival probability in CSCC patients and might be present in a small fraction ([~]20%) of advanced cancer patients. Collectively, these findings might enhance understanding of the CSCC immunological microenvironment and shed some light on the treatment of advanced CSCC.

cancer biology↗

bioRxiv2021

Opposing regulation of short-term memory by basal ganglia direct and indirect pathways that are coactive during behavior

The basal ganglia direct and indirect pathways are viewed to mediate opposing functions in movement. However, this classic model is challenged by recent findings that both pathways are coactive during behavior. We examined the roles of direct (dSPNs) and indirect (iSPNs) pathway spiny projection neurons in a decision-making task with a short-term memory (STM) component. Optogenetic stimulation of cortical-input-defined dSPNs and iSPNs during STM oppositely biased upcoming licking choice, without affecting licking execution. Optogenetically identified dSPNs and iSPNs showed similar response patterns, although with quantitative difference in spatiotemporal organization. To understand how coactive dSPNs and iSPNs play opposing roles, we recorded population activity in frontal cortex and the basal ganglia output nucleus SNr. Stimulation of dSPNs and iSPNs bidirectionally regulated cortical decision variable through the differential modulation of SNr ramping activity. These results reconcile different views by demonstrating that coactive dSPNs and iSPNs precisely shape cortical activity in a push-pull balance.

neuroscience↗

bioRxiv2021

Functional coupling between TRPV4 channel and TMEM16F modulates human trophoblast fusion

TMEM16F, a Ca2+-activated phospholipid scramblase (CaPLSase), is critical for placental trophoblast syncytialization, HIV infection, and SARS-CoV2-mediated syncytialization. How TMEM16F is activated during cell fusion is unclear. Here, we used trophoblasts as a model for cell fusion and demonstrated that Ca2+ influx through Ca2+ permeable transient receptor potential vanilloid channel TRPV4 is critical for TMEM16F activation and subsequent human trophoblast fusion. GSK1016790A, a TRPV4 specific agonist, robustly activates TMEM16F in trophoblasts. Patch-clamp electrophysiology demonstrated that TRPV4 and TMEM16F are functionally coupled within Ca2+ microdomains in human trophoblasts. Pharmacological inhibition or gene silencing of TRPV4 hindered TMEM16F activation and subsequent trophoblast syncytialization. Our study uncovers the functional expression of TRPV4 and a physiological activation mechanism of TMEM16F in human trophoblasts, thus providing us with novel strategies to regulate CaPLSase activity as a critical checkpoint of physiologically- and disease-relevant cell fusion events.

physiology↗

bioRxiv2021

Develop an efficient and specific AAV-based labeling system for Muller glia in mice

Cell degeneration in the retina leads to several ocular diseases and vision loss. Considerable research efforts focus on reprogramming Muller glia (MG) into functional cells to rescue vision as a promising therapeutic strategy, although whether MG can convert into functional bipolar cells, retinal ganglia cells (RGCs), rods or cones in mammals remains controversial. The broad applicability of tracking MG differentiation thus presents a need for improved labeling efficiency and specificity. Here, we compared AAV-based labeling strategies with conventional lineage-tracking by crossing transgenic mouse lines. We found that reporter expression was weak and not MG-specific in mGFAP-Cre transgenic mice. Different AAV serotypes showed a range of efficiency and specificity in labeling MG, leading us to optimize a human GFAP-Cre reporter system packaged in the AAV9 serotype with the WPRE (WPRE, woodchuck hepatitis virus post-transcriptional regulatory element) removed. The hGFAP-Cre- {Delta}WPRE reporter could label 20-73.8% MGs, with non-specific RGC labeling rates ranging from 0-0.08% at doses of 1 x 108 to 1010 vector genomes (vg) per eye, an approximate 40-fold reduction compared with the AAV9-hGFAP-Cre-WPRE labeling system. Furthermore, we validated the availability of the label system to trace MG reprogramming and also detected false-positive results reported previously. The AAV9-hGFAP-Cre-{Delta}WPRE system thus represents a highly efficient and specific labeling system for MG, providing a valuable tool for tracking cell fate in vivo.

neuroscience↗

bioRxiv2021

Cell free extrachromosomal circular DNA is common in human urine

Cell free extrachromosomal circular DNA (eccDNA) is evolving as a potential biomarker in liquid biopsies for disease diagnosis. In this study, an optimized next generation sequencing-based Circle-Seq method was developed to investigate urinary cell free eccDNA (ucf-eccDNA) from 28 adult healthy volunteers (mean age = 28, 19 males/ 9 females). The genomic distributions and sequence compositions of ucf-eccDNAs were comprehensively characterized. Approximately 1.2 million unique ucf-eccDNAs are identified, covering 14.9% of the human genome. Comprehensive characterization of ucf-eccDNAs show that ucf-eccDNAs contain higher GC content than flanking genomic regions. Most eccDNAs are less than 1000 bp and present four pronounced peaks at 203, 361, 550 and 728 bp, indicating the association between eccDNAs and the numbers of intact nucleosomes. Analysis of genomic distribution of ucf-eccDNAs show that eccDNAs are found in all chromosomes but enriched in chromosomes i.e. chr.17, 19 and 20 with high density of protein-codding genes, CpG islands, SINE and simple repeat elements. Lastly, analysis of sequence motif signatures at eccDNA junction sites reveal that direct repeats (DRs) are commonly found, indicating a potential role of DRs in eccDNA biogenesis. This work underscores the deep sequencing analysis of ucf-eccDNAs and provides a valuable reference resource for exploring potential applications of ucf-eccDNA as diagnostic biomarkers of urogenital disorders in the future. Significance StatementExtrachromosomal circular DNA (eccDNA) is an important genetic element and a biomarker for disease diagnosis and treatment. In this study, we conduct a comprehensive characterization of urinary cell free eccDNA (ucf-eccDNA) in 28 heathy subjects. Over one million ucf-eccDNAs are identified. Ucf-eccDNAs are characterized as high GC content. The size of most ucf-eccDNAs is less than 1000 bp and enriched in four peaks resembling the size of single, double, triple, and quadruple nucleosomes. The genomic distribution of ucf-eccDNAs is enriched in generic regions, protein-coding genes, Alu, CpG islands, SINE and simple repeats. Sequence motif analysis of ucf-eccDNA junctions identified simple direct repeats (DRs) commonly presented in most eccDNAs, suggesting potential roles of DRs in eccDNA biogenesis.

genomics↗

bioRxiv2021

Two decades of suspect evidence for adaptive molecular evolution - Negative selection confounding positive selection signals

There is a large literature in the last two decades affirming adaptive DNA sequences evolution between species. The main lines of evidence are from i) the McDonald-Kreitman (MK) test, which compares divergence and polymorphism data, and ii) the PAML test, which analyzes multi-species divergence data. Here, we apply these two tests concurrently on the genomic data of Drosophila and Arabidopsis. To our surprise, the >100 genes identified by the two tests do not overlap beyond random expectation. Because the non-concordance could be due to low powers leading to high false-negatives, we merge every 20 - 30 genes into a "supergene". At the supergene level, the power of detection is large but the calls still do not overlap. We rule out methodological reasons for the non-concordance. In particular, extensive simulations fail to find scenarios whereby positive selection can only be detected by either MK or PAML, but not both. Since molecular evolution is governed by positive and negative selection concurrently, a fundamental assumption for estimating one (say, positive selection) is that the other is constant. However, in a broad survey of primates, birds, Drosophila and Arabidopsis, we found that negative selection rarely stays constant for long in evolution. As a consequence, the variation in negative selection is often mis-construed as signals of positive selection. In conclusion, MK, PAML or any method that examines genomic sequence evolution has to explicitly address the variation in negative selection before estimating positive selection. In a companion study, we propose a possible path forward in two stages - first, by mapping out the changes in negative selection and then using this map to estimate positive selection. For now, the large literature on positive selection between species has to await the re-assessment.

evolutionary biology↗

bioRxiv2021

RPRM negatively regulates ATM levels involving its phosphorylation mediated by CDK4/CDK6

Sensitizing cancer cells to radio- and chemotherapy remains a hot topic in cancer treatment. Here it is identified that Protein Reprimo (RPRM) negatively regulates the levels of ataxia-telangiectasia mutated (ATM) protein kinase, a master regulator of DNA damage response (DDR) in the presence of DNA double-strand breaks (DSBs), resulting in impaired DNA repair efficiency and enhanced cellular sensitivity to genotoxic agents. Mechanistically, although RPRM is primarily located in cytoplasm, it rapidly translocates to nucleus shortly after induced by X-irradiation, interacts with ATM and promotes the nuclear export and proteasomal degradation of ATM. The nuclear translocation of RPRM is associated with its phosphorylation at serine 98, which is mediated by cyclin-dependent kinases 4/6 (CDK4/6). Inhibition of CDK4/6 stabilizes RPRM and promotes its nuclear import, in turn enhances the nuclear export of ATM and the reduction of ATM levels. As a result, RPRM overexpression and its phosphorylation inhibition sensitize cells to genotoxic agents. Moreover, RPRM deficiency significantly increases resistance to radiation-induced damage both in vitro and in vivo. These findings establish a crucial regulatory mechanism in which ATM is negatively modulated by RPRM, suggesting that RPRM may serve as a novel target for both cancer therapy and radiation protection.

cancer biology↗

bioRxiv2021

Efficient transformation and genome editing of watermelon assisted by genes that encode developmental regulators

AbstructThe Cucurbitaceae contains multiple species of important food plants. But most of them are difficult to be genetically transformed. Watermelon is one of the most important fruit species of Cucurbitaceae, and it is a model horticulture crops. Its low transgenic efficiency is the major bottleneck in functional genome research and genome editing-based breeding. Here, with the help of genes that encode developmental regulators (DRs), in particular AtGRF5, and an appropriate Agrobacterium strain (GV3101), we have significantly increased the transformation efficiency of watermelon to about 25%, which is a 40-fold increase compared with a traditional vector. AtGRF5-mediated transformation increased the number of transformable watermelon genotypes without causing obvious side effects. Moreover, when applying this strategy to CRISPR/Cas9-mediated genome editing, clpds knockout mutants were generated in the T0 generation. Our results show that AtGRF5 is a powerful and robust tool to effectively create transgenic plants or knockout mutations in watermelon. Similar strategies using DRs might also be able to overcome the transformation barriers in many other Cucurbitaceae species.

plant biology↗

bioRxiv2021

Diversity, function and evolution of aquatic vertebrate genomes

Aquatic vertebrates consist of jawed fish (cartilaginous fish and bony fish), aquatic mammals, reptiles and amphibians. Here, we present a comprehensive analysis of 630 aquatic vertebrate genomes to generate a standardized compendium of genomic data. We demonstrate its value by assessing their genome features as well as illuminating gene families related to the transition from water to land, such as Hox genes and olfactory receptor genes. We found that LINEs are the major transposable element (TE) type in cartilaginous fish and aquatic mammals, while DNA transposons are the dominate type in bony fish. To our surprise, TE types are not fixed in amphibians, the first group that transitioned to living on land. These results illustrate the value of a unified resource for comparative genomic analyses of aquatic vertebrates. Our data and strategy are likely to support all evolutionary and ecological research on vertebrates.

genomics↗

bioRxiv2021

Diversity, function and evolution of marine invertebrate genomes

Invertebrates, animals (metazoans) without backbones, encompass [~]97% of all animal yet remains understudied. They have provided insights into molecular mechanisms underlying fundamentally identical mechanisms in phylogenetically diverse animals, including vertebrates. Marine invertebrates have long fascinated researchers due to their abundance, diversity, adaptations, and impact on ecosystems and human economies. Here, we report a compendium and appraisal of 190 marine invertebrate genomes spanning 21 phyla, 43 classes, 92 orders, and 134 families. We identify a high proportion and long unit size of tandem repeats, likely contributing to reported difficulties in invertebrate genome assembly. A well-supported phylogenetic tree of marine invertebrates from 974 single-copy orthologous genes resolved topological controversies. We show that Ctenophora is at the basal phylum and Porifera is the sister group of Parahoxozoa; that Xenacoelomorpha is within Bilateria and is the sister group to Protostomia, rejecting three out of four hypotheses in the field; and that Bryozoa is at the basal position of Lophotrochozoa, not grouped into Lophophorata. We also present insights into the genetic underpinnings of metazoans from Hox genes, innate immune gene families, and nervous system gene families. Our marine invertebrate genome compendium provides a unified foundation for studies on their evolution and effects on ecological systems and human life.

genomics↗