Biology subjects

Zhou, M.

Publications and source records attributed to Zhou, M..

At least 145 records · Page 8Linked to original sources

bioRxiv2022

Lipoprotein sorting to the cell surface via a crosstalk between the Lpt and Lol pathways during outer membrane biogenesis

Lipopolysaccharide (LPS) and lipoprotein, two essential components of the outer membrane (OM) in Gram-negative bacteria, play critical roles in bacterial physiology and pathogenicity. LPS translocation to the OM is mediated by LptDE, yet how lipoproteins sort to the cell surface remains elusive. Here we report the identification of an inventory of lipoproteins that are transported to the cell surface via LptDE. Notably, we determined crystal structures of LptDE from Pseudomonas aeruginosa and its complex with an endogenous Escherichia coli lipoprotein YifL. The paLptDE-YifL structure demonstrates that YifL translocates to the OM via LptDE, in a manner similar to LPS transport. The {beta}-barrel domain serves as a passage for the proteinaceous moiety while its acyl chains are transported outside. Our finding has been corroborated by results from native mass spectrometry, immunofluorescence, and photocrosslinking assays, revealing a unique mechanism through which lipoproteins are translocated across the OM in an ATP- and LPS-dependent manner. Moreover, our study expands the scope of current knowledge of lipoprotein sorting by disclosing a crosstalk between the Lpt and Lol pathways.

microbiology↗

bioRxiv2022

Dynamic Interaction Learning and MultimodalRepresentation for Drug Response Prediction

Mining multimodal pharmaceutical data is crucial for in-silico drug candidate screening and discovery. A daunting challenge of integrating multimodal data is to enable dynamic feature modeling generalizable for real-world applications. Unlike conventional approaches using a simple concatenation with fixed parameters, in this paper, we develop a dynamic interaction learning network to adaptively integrate drug and different reactants on multimodal tasks towards robust drug response prediction. The primary objective of dynamic learning falls into two key aspects: at micro-level, we aim to dynamically search specific relational patterns on the whole reactant range for each drug-reactant pair; at macro-level, drug features can be used to adaptively correlate with different reactants. Extensive experiments demonstrate the validity of our approach in both drug protein interaction (DPI) and cancer drug response (CDR) tasks. Our approach achieves superior performance on both DPI (AUC = 0.967) and CDR (AUC = 0.932) tasks, outperforming competitive baselines from four real-world, drug-outcome datasets. In addition, the performance on the challenging blind subsets is remarkably improved, where AUC value increases from 0.843 to 0.937 on blind protein set of DPI task, and Pearsons correlation value increases from 0.516 to 0.566 on blind drug set of CDR task. A series of case studies highlight the potential generalization and interpretability of dynamic learning in the in-silico drug response assessment.

bioinformatics↗

bioRxiv2022

Caught in the Act: targeted mutagenesis of the herpesvirus glycoprotein B central helix captures fusion transition states

Herpesviruses remain a burden for animal and human health, including the medically important varicella-zoster virus (VZV). Membrane fusion mediated by conserved core glycoproteins, the fusogen gB and the heterodimer gH-gL, enables herpesvirus cell entry. The ectodomain of gB orthologs has five domains and is proposed to transition from a prefusion to postfusion conformation but the functional relevance of the domains for this transition remains poorly defined. Structure-function studies of the VZV gB DIII central helix were performed targeting residues 526EHV528. Critically, a H527P mutation captured gB in a prefusion conformation as determined by cryo-EM, a loss of membrane fusion in a virus free assay, and failure of recombinant VZV to spread in cell monolayers. Importantly, two predominant cryo-EM structures of gB[H527P] were identified by 3D classification and focused refinement, suggesting they represented gB conformations in transition. These studies reveal gB DIII as a critical element for herpesvirus gB fusion function.

microbiology↗

bioRxiv2022

Single-molecule long-read methylation profiling reveals regional DNA methylation regulated by Elongator Complex Subunit 2 in Arabidopsis roots experiencing spaceflight

BackgroundThe Advanced Plant Experiment-04 - Epigenetic Expression (APEX04-EpEx) experiment onboard the International Space Station examined the spaceflight-altered cytosine methylation in two genetic lines of Arabidopsis thaliana, wild-type Col-0 and the mutant elp2-5, which is deficient in an epigenetic regulator Elongator Complex Subunit 2 (ELP2). Whole-genome bisulfite sequencing (WGBS) revealed distinct spaceflight associated methylation differences, presenting the need to explore specific space- altered methylation at single-molecule resolution to associate specific changes over large regions of spaceflight related genes. To date, tools of multiplexed targeted DNA methylation sequencing remain limited for plant genomes. ResultsTo provide methylation data at single-molecule resolution, Flap-enabled next-generation capture (FENGC), a novel targeted multiplexed DNA capture and enrichment technique allowing cleavage at any specified sites, was applied to survey spaceflight-altered DNA methylation in genic regions of interest. The FENGC capture panel contained 108 targets ranging from 509 to 704 nt within the promoter or gene body regions of gene targets derived from spaceflight whole-genome data sets. In addition to genes with significant changes in expression and average methylation levels between spaceflight and ground control, targets with space-altered distributions of the proportion of methylated cytosines per molecule were identified. Moreover, trends of co-methylation of different cytosine contexts were exhibited in the same DNA molecules. We further identified significant DNA methylation changes in three previously biological process- unknown genes, and loss-of-function mutants of two of these genes (named as EMO1 and EMO2 for ELP2-regulated Methylation in Orbit 1 and 2) showed enhanced root growth rate. ConclusionsFENGC simplifies and reduces the cost of multiplexed, targeted, single-molecule profiling of methylation in plants, providing additional resolution along each DNA molecule that is not seen in population-based short-read data such as WGBS. This case study has revealed spaceflight-altered regional modification of cytosine methylation occurring within single DNA molecules of cell subpopulations, which were not identified by WGBS. The single-molecule survey by FENGC can lead to identification of novel functional genes. The newly identified EMO1 and EMO2 are root growth regulators which may be epigenetically involved in plant adaptation to spaceflight.

plant biology↗

bioRxiv2022

Flap-enabled next-generation capture (FENGC): precision targeted single-molecule profiling of epigenetic heterogeneity, chromatin dynamics, and genetic variation

Targeted sequencing is an increasingly sought technology. Available methods, however, are often costly and yield high proportions of off-target reads. Here, we present FENGC, a scalable, multiplexed method in which target sequences are assembled into 5' flaps for precise excision by flap endonuclease. Recovery of length-matched sequences, amplification with universal primers, and exonucleolytic removal of non-targeted genomic regions mitigate amplification biases and consistently yield [≥] 80% on-target sequencing. Furthermore, optimized sequential reagent addition and purifications minimize sample loss and facilitate rapid processing of sub-microgram quantities of DNA for detection of genetic variants and DNA methylation. Treatment of cultured human glioblastoma cells and primary murine monocytes with GC methyltransferase followed by FENGC and high-coverage enzymatic methyl sequencing provides single-molecule, long-read detection of differential endogenous CG methylation, dynamic nucleosome repositioning, and transcription factor binding. FENGC provides a versatile and cost-effective platform for targeted sequence enrichment for analysis of genetic and/or epigenetic heterogeneity.

cancer biology↗

bioRxiv2022

Universal chimeric Fcγ receptor T cells with appropriate affinity for IgG1 antibody exhibit optimal antitumor efficacy

Developing universal CARs with improved flexible targeting and controllable activities is urgently needed. While several studies have suggested the potential of CD16a in tandem with monoclonal antibodies to construct universal CAR T cells, the weak affinity between them is one of the limiting factors for efficacy. Herein, we systematically investigated the impact of Fc{gamma} receptor (Fc{gamma}R) affinity on CAR T cells properties by constructing universal CARs using Fc{gamma} receptors with different affinities for IgG1 antibodies, namely CD16a, CD32a, and CD64. We demonstrated that the activities of these universal CAR T cells on tumor cells could be redirected and regulated by IgG1 antibodies. In xenografted mice, 64CAR chimeric Jurkat cells with the highest affinity showed significant antitumor effects in combination with herceptin in the Her2 low expression U251 MG model. However, in the CD20 high expression Raji model, 64CAR caused excessive activation of CAR-T cells, which resulted in cytokine release syndrome (CRS) and the decline of antitumor activity, and 32CAR with a moderate affinity brought the best efficacy. Our work extended the knowledge about Fc{gamma}R-based universal CAR T cells and suggested that only the Fc{gamma}RCAR with an appropriate affinity can offer the optimal antitumor advantages of CAR T cells. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC="FIGDIR/small/513394v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1348ceorg.highwire.dtl.DTLVardef@10eb32org.highwire.dtl.DTLVardef@c12aeforg.highwire.dtl.DTLVardef@1e9c9ea_HPS_FORMAT_FIGEXP M_FIG C_FIG Universal CAR T cells based on Fc{gamma} receptors exhibit a specific tumor-killing effect. However, the affinities of Fc{gamma} receptors greatly influence the efficacy and adverse effects in vivo.

immunology↗

bioRxiv2022

Intranasal delivery of NS1-deleted influenza virus vectored COVID-19 vaccine restrains the SARS-CoV-2 inflammatory response

The emergence of SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus-2) variants and "anatomical escape" characteristics threaten the effectiveness of current coronavirus disease (COVID-19) vaccines. There is an urgent need to understand the immunological mechanism of broad-spectrum respiratory tract protection to guide broader vaccines development. In this study, we investigated immune responses induced by an NS1-deleted influenza virus vectored intranasal COVID-19 vaccine (dNS1-RBD) which provides broad-spectrum protection against SARS-CoV-2 variants. Intranasal delivery of dNS1-RBD induced innate immunity, trained immunity and tissue-resident memory T cells covering the upper and lower respiratory tract. It restrained the inflammatory response by suppressing early phase viral load post SARS-CoV-2 challenge and attenuating pro-inflammatory cytokine (IL-6, IL-1B, and IFN-{gamma}) levels, thereby reducing excess immune-induced tissue injury compared with the control group. By inducing local cellular immunity and trained immunity, intranasal delivery of NS1-deleted influenza virus vectored vaccine represents a broad-spectrum COVID-19 vaccine strategy to reduce disease burden.

immunology↗

bioRxiv2022

MAM-STAT3-induced upregulation of mitochondrial Ca+2 causes immunosenescence in patients with type A mandibuloacral dysplasia

Homozygous lamina/c p.R527C mutations result in severe mandibuloacral dysplasia (MAD) and progeroid syndrome, but the underlying molecular pathology remains unknown. Here, we report on three patients with MAD, all displaying severe systemic inflammaging and characterized the major molecular pathways involved in the manifestation of this disease. Analysis of induced pluripotent stem cell (IPSC)-derived mesenchymal stem cells (MAD-iMSCs) obtained from the patients revealed that increased mitochondrial Ca+2 loading was the root cause of lost mitochondrial membrane potential, abnormal fission/fusion and fragmentation, which then participated in inflammaging by inducing the inflammasome. These alterations in Ca+2 homeostasis were mediated by signal transducer and activator of transcription 3 (STAT3), which is located on the mitochondrial associated membrane (MAM). STAT3 function could be rescued by treatment with clinically-approved IL-6 blockers, or by correction of R527C mutations. In addition, extracellular vesicles (EVs) obtained from MAD-iMSCs displayed reduced immunomodulatory function, being unable to rescue bleomycin-induced lung fibrosis and triggering mitochondrial dysfunction, senescence, and fibrosis in healthy cells. Our results provide new insights into the pathology of complex lamin-associated MAD with systemic immunosenescence, and suggest that targeting defective mitochondrial Ca+2 homeostasis may represent a promising novel therapy for this condition.

cell biology↗

bioRxiv2022

Mechanism of Ca2+ transport by ferroportin

Ferroportin (Fpn) is a transporter that releases Fe2+ from cells and is important for iron homeostasis in circulation. Export of Fe2+ by Fpn is coupled to import of H+ to maintain charge balance. Although Ca2+ was shown to modulate Fe2+ transport in Fpn, transport of Ca2+ by Fpn has not been demonstrated. Here we show that human Fpn (HsFpn) mediates Ca2+ transport, and that the Ca2+ transport does not rely on the transport of other ions. We determine the structure of Ca2+-bound HsFpn and identify a single Ca2+ binding site distinct from the Fe2+ binding sites. Further studies validate the Ca2+ binding site and show that Ca2+ transport is inhibited in the presence of Fe2+ but not vice versa. Function of Fpn as a Ca2+ uniporter in the absence of Fe2+ provides a molecular basis for regulations of iron homeostasis by Ca2+.

biochemistry↗

bioRxiv2022

Structural basis of ferroportin inhibition by minihepcidin PR73

Ferroportin (Fpn) is the only known iron exporter in humans and is essential for maintaining iron homeostasis. Fpn activity is suppressed by hepcidin, an endogenous peptide hormone, which inhibits iron export and promotes endocytosis of Fpn. Hepcidin deficiency leads to hemochromatosis and iron-loading anemia. Previous studies have shown that small peptides that mimic the first few residues of hepcidin, i.e. minihepcidins, are more potent than hepcidin. However, the mechanism of enhanced inhibition by minihepcidins remains unclear. Here, we report the structure of human ferroportin in complex with a minihepcidin, PR73 that mimics the first 9 residues of hepcidin, at 2.7 [A] overall resolution. The structure reveals novel interactions that were not present between Fpn and hepcidin. We validate PR73-Fpn interactions through binding and transport assays. These results provide insights into how minihepcidins increase inhibition potency and will guide future developments of Fpn inhibitors.

biochemistry↗

bioRxiv2022

BCAA catabolism drives adipogenesis via an intermediate metabolite and promotes subcutaneous adipose tissue expansion during obesity

Branched-chain amino acids (BCAAs, including leucine, isoleucine, and valine) have emerged as major players in metabolic health and diseases, but the underlying mechanisms remain obscure. Here, we report that BCAA catabolism drives adipogenesis via an intermediate metabolite of leucine and promotes subcutaneous white adipose tissue (sWAT) expansion during obesity. Genetic analyses of humans and mice reveal that the BCAA catabolic pathway in WAT is strongly correlated with adipose physiology and obesity traits. Altering BCAA catabolism in mature adipocytes exerts minor effects on adiposity in mice. However, enhancing adipose BCAA catabolism via FABP4-Cre-mediated Bckdk deletion promotes diet-induced obesity while blocking adipose BCAA catabolism through Bckdha ablation does the opposite. Intriguingly, the catabolism of BCAAs elicits fat depot-specific responses and promotes sWAT extension and adipogenesis in a cell-autonomous manner. Mechanistically, BCAA catabolism drives adipocyte differentiation via an intermediate metabolite of leucine, which activates mTORC1 and polyamine synthesis from methionine to promote the expression of adipogenic master regulators. Together, these results demonstrate that BCAA catabolism promotes adipogenesis and sWAT expansion during obesity. The crosstalk between leucine and methionine metabolism driven by the catabolic intermediate highlights an unexpected regulatory role of amino acids in metabolic health and diseases.

physiology↗

bioRxiv2022

Strigolactone mediates moso bamboo root response to phosphate stress

Moso bamboo (Phyllostachys edulis) grows in soils with widespread phosphate (Pi) deficiency, resulting in low shoot yield. While it also suffers from high Pi due to heavy fertilization, which causes the degradation of bamboo forest. A novel plant hormone, strigolactone (SL), plays a key role in root growth and development under Pi stress, but its regulatory mechanism has not been systematically reported. Our study investigated SL-mediated mechanism in response to Pi stress in moso bamboo. We compared the root growth under low, sufficient and high Pi, and analyzed the temporary trends of gene expression in primary root tip region and lateral root primordium zone. The effects of SL-analog (GR24) and SL-inhibitor (TIS108) on root architecture at low and high Pi were evaluated. SL biosynthesis and signaling are the main pathways of root response to Pi stress. With the decrease of Pi level, 5-deoxystrigol and strigol in the root exudates increase significantly. Under low Pi, SL is indispensable for the maintenance of root cell morphology, and promotes primary root elongation and reduces lateral root formation by upregulating the expression of phosphorous starvation response genes and downregulating the expression of abscisic acid response genes. The absence of SL at high Pi releases the inhibition of ethylene responsive genes expression to inhibit root elongation and promote branching. In general, SL mediates the response of bamboo roots to Pi stress by regulating its biosynthesis and signal transduction and influencing other hormone pathways.

molecular biology↗

bioRxiv2022

Deep generative modeling and clustering of single cell Hi-C data

Deciphering 3D genome conformation is important for understanding gene regulation and cellular function at a spatial level. The recent advances of single cell Hi-C technologies have enabled the profiling of the 3D architecture of DNA within individual cell, which allows us to study the cell-to-cell variability of 3D chromatin organization. Computational approaches are in urgent need to comprehensively analyze the sparse and heterogeneous single cell Hi-C data. Here, we proposed scDEC-Hi-C, a new framework for single cell Hi-C analysis with deep generative neural networks. scDEC-Hi-C outperforms existing methods in terms of single cell Hi-C data clustering and imputation. Moreover, the generative power of scDEC-Hi-C could help unveil the heterogeneity of chromatin architecture across different cell types. We expect that scDEC-Hi-C could shed light on deepening our understanding of the complex mechanism underlying the formation of chromatin contacts. scDEC-Hi-C is freely available at https://github.com/kimmo1019/scDEC-Hi-C. Key pointsO_LIscDEC-Hi-C provides an end-to-end framework based on autoencoder and deep generative model to comprehensively analyze single cell Hi-C data, including low-dimensional embedding and clustering. C_LIO_LIThrough a series of experiments including single cell Hi-C data clustering and structural difference identification, scDEC-Hi-C demonstrates suprioir performance over existing methods. C_LIO_LIIn the downstream analysis of chromatin loops from single cell Hi-C data, scDEC-Hi-C is capable of significantly enhancing the ability for identifying single cell chromatin loops by data imputation. C_LI

bioinformatics↗

bioRxiv2022

Comprehensive analysis identifies histone 2A ubiquitination as a marker for aging and pan-cancer prognosis

Ubiquitination is a post-translational modification (PTM) that induces protein degradation or function alteration and plays crucial roles in aging and cancer. Previous ubiquitinomes of aging mainly focused on how ubiquitination changes in drosophila and other lower animals, but how ubiquitination changes during the aging of higher animals and what causes these changes remain unclear. Here, we profiled whole-life ubiquitinome data of mouse brain, heart, liver, muscle, and spleen, and integratively analyzed the ubiquitinome data with RNA sequencing data. The results showed that the ubiquitination of protein, especially histone 2A (H2A), changed intensely during aging due to the regulated expression of E3 ligases (E3s) and deubiquitylating enzymes (DUBs). Then we developed two distinct H2A E3s/DUBs expression subtypes with different prognosis, DNA damage response (DDR), and tumor microenvironment cell infiltration degrees based on an unsupervised method in pan-cancer. In conclusion, our study provided temporal resolution ubiquitinome data of mouse aging and revealed the vital role of H2A ubiquitination in aging and tumor progression.

cancer biology↗

bioRxiv2022

BayesTME: A unified statistical framework for spatial transcriptomics

Spatial variation in cellular phenotypes underlies heterogeneity in immune recognition and response to therapy in cancer and many other diseases. Spatial transcriptomics (ST) holds the potential to quantify such variation, but existing analysis methods address only a small part of the analysis challenge, such as spot deconvolution or spatial differential expression. We present BayesTME, an end-to-end Bayesian method for analyzing spatial transcriptomics data. BayesTME unifies several previously distinct analysis goals under a single, holistic generative model. This unified approach enables BayesTME to (i) be entirely reference-free without any need for paired scRNA-seq, (ii) outperform a large suite of methods in quantitative benchmarks, and (iii) uncover a new type of ST signal: spatial differential expression within individual cell types. To achieve the latter, BayesTME models each phenotype as spatially adaptive and discovers statistically significant spatial patterns amongst coordinated subsets of genes within phenotypes, which we term spatial transcriptional programs. On human and zebrafish melanoma tissues, BayesTME identifies spatial transcriptional programs that capture fundamental biological phenomena like bilateral symmetry, differential expression between interior and surface tumor cells, and tumor-associated fibroblast and macrophage reprogramming. Our results demonstrate BayesTMEs power in unlocking a new level of insight from spatial transcriptomics data and fostering a deeper understanding of the spatial architecture of the tumor microenvironment. BayesTME is open source and publicly available (https://github.com/tansey-lab/bayestme).

bioinformatics↗

bioRxiv2022

Cryo-EM structures reveal a dynamic transformation process of human alpha-2-macroglobulin working as a protease inhibitor

Human alpha-2-macroglobulin is a well-known proteases inhibitor against a broad spectrum of proteases. It also plays important roles in immunity, inflammation, and infections. Here, we report cryo-EM structures of human alpha-2-macroglobulin of the native state, the transformed state induced by its authentic substrate, human trypsin, and serial intermediate states between the native and the fully induced state. These structures exhibit distinct conformations, which reveal a dynamic transformation process of alpha-2-macroglobulin acting as a protease inhibitor. The results shed light on the molecular mechanism of alpha-2-macroglobulin entrapping substrates, and help to understand how alpha-2-macroglobulin possesses variant physiological functions.

biochemistry↗

bioRxiv2022

Structure Of Eukaryotic Type III Glutamine Synthetase Lacking Ring-Ring Quaternary Contacts

There are three major classes of the enzyme Glutamine Synthetase (GS), denoted as GSI, GSII and GSIII in both prokaryotes and eukaryotes, that share widely conserved active-site residues but vary in average protein length, oligomeric assembly state and cofactor requirements. Key structures have been determined for all prokaryotic and eukaryotic GS classes except the GSIII class of eukaryotic origin, where even basic questions about its functional oligomeric state are unresolved. Here, we report the 3.2[A] cryo-EM structure of eukaryotic GSIII from the ancient organism of green algal lineage Ostreococcus tauri. Using a combined structural and biochemical approach, we demonstrate that O. tauri GSIII self-assembles and functions exclusively as a single hexameric ring. This is unlike most GSs in other classes, where they form double-stacked ring assemblies instead. Major structural differences in the ring-ring stacking across the classes suggest evolutionary pressure may have favored higher-order interactions that might be beneficial but are not necessarily required for the catalytic GS performance.

biochemistry↗

bioRxiv2022

Cryo-EM structure of the diapause chaperone artemin

The protein artemin constitutes over 10% of all protein in Artemia cysts during diapause and acts as both an RNA and protein chaperone. However, its mechanistic details remain elusive since no high-resolution structure of artemin exists. Here we report the full-length structure of artemin at 2.04 [A] resolution. The cryo-EM map contains density for an intramolecular disulfide bond between Cys22-Cys61 and resolves the entire C-terminus extending into the core of the assembled protein cage. We also provide data supporting the role of C-terminal helix F towards stabilizing the dimer form that is believed to be important for its chaperoning activity. We were able to destabilize this effect by placing a tag at the C-terminus to fully pack the internal cavity and cause limited steric hindrance.

biochemistry↗