Biology subjects

Zhao, J.

Publications and source records attributed to Zhao, J..

At least 325 records · Page 18Linked to original sources

bioRxiv2023

Biodecomposing Spirulina platensis by a de novo designed Bacillus subtilis-based method to develop a medium for the high cell-density cultivation of Escherichia coli in batch mode

High cell-density cultivation (HCDC) is fundamental to basic research and industrial applications, especially in batch mode. However, limited media are available for batch culture of HCDC, because the media for batch culture must contain extremely sufficient nutrients on the one hand and few or even no substrates to generate detrimental metabolite on the other hand to attain HCDC. Spirulina platensis (SP), a new media material, is considered ideal for the development of such media. Here, we develop a biolysis method for SP degradation based on the cultivation supernatant of Bacillus subtilis and extensively demonstrate its higher degradation (indicated by the production of more small peptides and free amino acids) and cultivation effectiveness with three other methods. Based on its SP hydrolysates, a modified SP-based broth (MSP) is then formulated. Tests on Escherichia coli K-12 show that MSP achieves HCDC with several benefits: (i) its maximum optical density at 600 nm is [~]16.67, significantly higher than that of Luria-Bertani (LB) broth (only [~]6.30); (ii) MSP requires only 36 h to reach peak growth, much faster than that of LB (48 h); (iii) its maximum growth (1.12 {+/-} 0.01 h-1) is significantly higher than that of LB (0.20 {+/-} 0.00 h-1); (v) MSP initiates growth immediately after inoculation (lag time <0), comparable to LB; (iv) the number of viable cells in MSP is high ([~]2.16 x 1011 ml-1), [~]10.19 times the amount in LB. Consequently, we envision MSP will become the first choice for E. coli HCDC batch culture in the future. ImportanceSo far, it is the first time to develop a high-efficiency method for transforming Spirulina platensis (SP) into medium ingredients. Based on its SP hydrolysates, a high cell-density cultivation (HCDC) medium for the batch culture of Escherichia coli is formulated for the first time, which is greatly beneficial for both basic research and industrial applications. In addition to HCDC, the SP hydrolysates can be extended to a wide range of applications, due to their rich nutrient content. Besides, this study demonstrates for the first time that SP is an ideal material to develop HCDC media. Furthermore, this study demonstrates that medium development and modification for batch culture can attain HCDC, without the development of new culture technologies. Therefore, this study highlights the importance of the rebirth of medium development and modification and supports the shift from developing new culture technologies to medium development and modification for HCDC in batch mode.

microbiology↗

bioRxiv2023

Trained immunity in human monocyte enhances myeloid-T-cell pathogenic crosstalk in Ankylosing Spondylitis

Recent studies in infectious, cardiovascular and neurodegenerative diseases have established the presence of memory in innate immune cells. This "trained immunity (TI)" leads to an enhanced response to a second challenge. Monocytes in Ankylosing Spondylitis (AS), a common form of inflammatory arthritis, are known to be hyper-responsive to microbial stimulus lipopolysaccharide (LPS). We asked if TI is present in AS monocytes and, if so, how it contributes to disease pathology. Using Single-cell RNA sequencing (scRNA-seq), flow cytometry and enzyme-linked immunosorbent assays (ELISA), we identify a subset of monocytes from AS patients exhibiting features of trained immunity and being hyperresponsive to LPS stimulation. Surprisingly, both trained monocytes in AS and {beta}-glucan-trained monocytes from healthy donors are hyper-responsive to T-cell-induced activation. scRNA-seq of AS synovial mononuclear cells shows enrichment of a monocyte population with these/analogous features. Additionally, T cell-stimulated monocytes act back on T-cells to support Th17 responses (of established pathology in AS). Lastly, using genetic and chemical perturbations we show that ERN1, an AS risk gene enriched in this trained monocyte population, contributes to T-cell-induced monocyte activation. Our data provide strong evidence for the first time for the key role of TI in common human inflammatory arthritis.

molecular biology↗

bioRxiv2023

ATR kinase supports normal proliferation in the early S phase by preventing replication resource exhaustion.

The ATR kinase, which coordinates cellular responses to DNA replication stress, is also essential for the proliferation of normal unstressed cells. Although its role in the replication stress response is well defined, the mechanisms by which ATR supports normal cell proliferation remain elusive. Here, we show that ATR is dispensable for the viability of G0-arrested naive B cells. However, upon cytokine-induced proliferation, Atr-deficient B cells initiate DNA replication efficiently in early S phase, but by mid-S phase they display dNTP depletion, fork stalling, and replication failure. Nonetheless, productive DNA replication can be restored in Atr-deficient cells by pathways that suppress origin firing, such as downregulation of CDC7 and CDK1 kinase activities. Together, these findings indicate that ATR supports the proliferation of normal unstressed cells by tempering the pace of origin firing during the early S phase to avoid exhaustion of dNTPs and other replication factors.

genetics↗

bioRxiv2023

Tumor-specific neutrophils originating from meninges promote glioblastoma

Glioma is one of the most aggressive human cancers with limited therapeutic options. Though research has extensively examined immune components in those malignant tumors, the pathophysiological mechanism establishing their immunosuppressive microenvironment remains incompletely characterized. In this study, we report for the first time the unique presence of tumor-specific neutrophils (TSNs) in human glioblastoma (GBM) tumors. This newly defined neutrophil subtype exhibits the high expression of several immunosuppressive genes (e.g., CD274 and IDO1) and is strongly correlated with glioma grades and poor prognosis of patients. TSNs with comparable gene signatures are similarly present in the tumors but not bone marrow or spleen of mouse glioma models. Blockage of TSN recruitment by either Cxcl1-knockout in glioma cells or Cxcr2-deletion in host mice significantly enhances antitumor immunity and inhibits tumor progression. Surprisingly, we further identify the meninges as the key extratumoral source of generating TSNs in both human GBM patients and mouse glioma models. These results have elucidated a novel mechanism of neutrophils designating the tumor immune microenvironment and the essential link of meningeal immunity to glioma.

cancer biology↗

bioRxiv2023

StressME: unified computing framework of Escherichia coli metabolism, gene expression, and stress responses

Generalist microbes have adapted to a multitude of environmental stresses through their integrated stress response system. Individual stress responses have been quantified by E. coli metabolism and expression (ME) models under thermal, oxidative and acid stress, respectively. However, the systematic quantification of crosstalk among these stress responses remains lacking. Here, we present StressME: the unified stress response model of E. coli combining thermal (FoldME), oxidative (OxidizeME) and acid (AcidifyME) stress responses. StressME is the most up to date ME model for E. coli and it reproduces all published single-stress ME models. Additionally, it includes refined rate constants to improve prediction accuracy for wild-type and stress-evolved strains. StressME revealed certain optimal proteome allocation strategies associated with cross-stress responses. These stress-optimal proteomes were shaped by trade-offs between protective vs. metabolic enzymes; cytoplasmic vs. periplasmic chaperones; and expression of stress-specific proteins. As StressME is tuned to compute metabolic and gene expression responses under mild acid, oxidative, and thermal stresses, it is useful for engineering and health applications. The modular design of our open-source package also facilitates model expansion (e.g., to new stress mechanisms) by the computational biology community. Author summaryA fundamental understanding of multi-stress adaptation in E.coli has potential industrial relevance. While individual stress responses have been quantified through the protein regulatory network in E.coli, the systematic quantification of the crosstalk among stress responses remains lacking. Here, we develop a new modeling pipeline by which thermal, oxidative and acid stress response can be coupled to each other, and the metabolic activities, protein and metabolic flux redistribution due to cross stress can be quantified. We optimize the effective rate constants in the integrated model. We then confirm the model robustness by validating against the published data under single stress. Finally, we use the model to characterize the cross-adaptation between protective and catalytic proteins as well as between chaperones present in different cellular compartments. We find effective cross-protection against cross stress by adapting the E.coli cells to the thermal stress first. We also indicate the presence of trade-offs by which the cell may refuse to give up more protein allocation away from one stress response to the other, because doing so would decrease stress tolerance further. The single stress plug-in design makes the model build-up pipeline flexible and expandable, allowing incorporation of more stressors into the model architecture for industrial applications.

systems biology↗

bioRxiv2023

TREM2 promotes glioma progression and angiogenesis mediated by microglia/brain macrophages

TREM2, a myeloid cell-specific signaling molecule, controls essential functions of microglia and impacts on the pathogenesis of Alzheimers disease and other neurodegenerative disorders. TREM2 is also highly expressed in tumor-associated macrophages and plays detrimental roles in an experimental mouse sarcoma model. Here we studied whether TREM2 influences glioma progression. We found a sex- dependent effect of TREM2: the glioma volume is significantly attenuated in TREM2- deficient male but not female mice injected with GL261-EGFP glioma cells. The accumulation of glioma-associated microglia/macrophages (GAMs) and vascularization is reduced in male TREM2-deficient mice. A transcriptomic analysis of glioma tissue revealed that TREM2 deficiency suppresses angiogenic genes and MHC clusters. In an organotypic slice model devoid of functional vascularization, the tumor size was not affected by TREM2-deficiency. In human resection samples from glioblastoma, TREM2 is upregulated in GAMs. Based on the TCGA and CGGA databases, the TREM2 expression levels are negatively correlated with survival. Thus, the TREM2-dependent crosstalk between GAMs and the vasculature formation promotes glioma growth. Graphic abstractTREM2-dependent crosstalk between glioma-associated microglia/macrophages and the vasculature formation promotes glioma growth in male glioma mouse model. Created with BioRender.com O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=137 SRC="FIGDIR/small/540621v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@65bec7org.highwire.dtl.DTLVardef@1d00961org.highwire.dtl.DTLVardef@117ee5eorg.highwire.dtl.DTLVardef@227994_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2023

Lipid-anchored Proteasomes Control Membrane Protein Homeostasis

Protein degradation in eukaryotic cells is mainly carried out by the 26S proteasome, a macromolecular complex not only present in the cytosol and nucleus but also associated with various membranes. How proteasomes are anchored to the membrane and the biological meaning thereof have been largely unknown in higher organisms. Here we show that N-myristoylation of the Rpt2 subunit is a general mechanism for proteasome-membrane interaction. Loss of this modification in the Rpt2-G2A mutant cells leads to profound changes in the membrane-associated proteome, perturbs the endomembrane system and undermines critical cellular processes such as cell adhesion, endoplasmic reticulum-associated degradation (ERAD) and membrane protein trafficking. Rpt2G2A/G2A homozygous mutation is embryonic lethal in mice and is sufficient to abolish tumor growth in a nude mice xenograft model. These findings have defined an evolutionarily conserved mechanism for maintaining membrane protein homeostasis and underscored the significance of compartmentalized protein degradation by myristoyl-anchored proteasomes (MAPs) in health and disease.

biochemistry↗

bioRxiv2023

Conformational dynamics of the μ-opioid receptor determine ligand intrinsic efficacy

The -opioid receptor (OR) is an important target for pain management and the molecular understanding of drug action will facilitate the development of better therapeutics. Here we show, using double electron-electron resonance (DEER) and single-molecule fluorescence resonance energy transfer (smFRET), how ligand-specific conformational changes of the OR translate into a broad range of intrinsic efficacies at the transducer level. We identify several cytoplasmic receptor conformations interconverting on different timescales, including a pre-activated receptor conformation which is capable of G protein binding, and a fully activated conformation which dramatically lowers GDP affinity within the ternary complex. Interaction of {beta}-arrestin-1 with the OR core binding site appears less specific and occurs with much lower affinity than binding of G protein Gi. One-Sentence SummaryLigand-dependent conformational dynamics of the -opioid receptor determine downstream signaling efficacy.

biochemistry↗

bioRxiv2023

A transposase-derived gene required for human brain development

DNA transposable elements and transposase-derived genes are present in most living organisms, including vertebrates, but their function is largely unknown. PiggyBac Transposable Element Derived 5 (PGBD5) is an evolutionarily conserved vertebrate DNA transposase-derived gene with retained nuclease activity in human cells. Vertebrate brain development is known to be associated with prominent neuronal cell death and DNA breaks, but their causes and functions are not well understood. Here, we show that PGBD5 contributes to normal brain development in mice and humans, where its deficiency causes disorder of intellectual disability, movement, and seizures. In mice, Pgbd5 is required for the developmental induction of post-mitotic DNA breaks and recurrent somatic genome rearrangements. In the brain cortex, loss of Pgbd5 leads to aberrant differentiation and gene expression of distinct neuronal populations, including specific types of glutamatergic neurons, which explains the features of PGBD5 deficiency in humans. Thus, PGBD5 might be a transposase-derived enzyme required for brain development in mammals. One-Sentence SummaryPiggyBac Transposable Element Derived 5 (PGBD5) is required for brain development in humans and mice through genetic and epigenetic mechanisms.

neuroscience↗

bioRxiv2023

3D structural human interactome reveals proteome-wide perturbations by disease mutations

Human genome sequencing studies have identified numerous loci associated with complex diseases. However, translating human genetic and genomic findings to disease pathobiology and therapeutic discovery remains a major challenge at multiscale interactome network levels. Here, we present a deep-learning-based ensemble framework, termed PIONEER (Protein-protein InteractiOn iNtErfacE pRediction), that accurately predicts protein binding partner-specific interfaces for all known protein interactions in humans and seven other common model organisms, generating comprehensive structurally-informed protein interactomes. We demonstrate that PIONEER outperforms existing state-of-the-art methods. We further systematically validated PIONEER predictions experimentally through generating 2,395 mutations and testing their impact on 6,754 mutation-interaction pairs, confirming the high quality and validity of PIONEER predictions. We show that disease-associated mutations are enriched in PIONEER-predicted protein-protein interfaces after mapping mutations from [~]60,000 germline exomes and [~]36,000 somatic genomes. We identify 586 significant protein-protein interactions (PPIs) enriched with PIONEER-predicted interface somatic mutations (termed oncoPPIs) from pan-cancer analysis of [~]11,000 tumor whole-exomes across 33 cancer types. We show that PIONEER-predicted oncoPPIs are significantly associated with patient survival and drug responses from both cancer cell lines and patient-derived xenograft mouse models. We identify a landscape of PPI-perturbing tumor alleles upon ubiquitination by E3 ligases, and we experimentally validate the tumorigenic KEAP1-NRF2 interface mutation p.Thr80Lys in non-small cell lung cancer. We show that PIONEER-predicted PPI-perturbing alleles alter protein abundance and correlates with drug responses and patient survival in colon and uterine cancers as demonstrated by proteogenomic data from the National Cancer Institutes Clinical Proteomic Tumor Analysis Consortium. PIONEER, implemented as both a web server platform and a software package, identifies functional consequences of disease-associated alleles and offers a deep learning tool for precision medicine at multiscale interactome network levels.

bioinformatics↗

bioRxiv2023

Contigs directed gene annotation (ConDiGA) for accurate protein sequence database construction in metaproteomics

Microbiota are closely associated to human health and disease. Metaproteomics can provide a direct means to identify microbial proteins in microbiota for compositional and functional characterization. However, in-depth and accurate metaproteomics is still limited due to the extreme complexity and high diversity of microbiota samples. One of the main challenges is constructing a protein sequence database that best fits the microbiota sample. Herein, we proposed an accurate taxonomic annotation pipeline from metagenomic data for deep metaproteomic coverage, namely contigs directed gene annotation (ConDiGA). We mixed 12 known bacterial species to derive a synthetic microbial community to benchmark metagenomic and metaproteomic pipelines. With the optimized taxonomic annotation strategy by ConDiGA, we built a protein sequence database from the metagenomic data for metaproteomic analysis and identified about 12,000 protein groups, which was very close to the result obtained with the reference proteome protein sequence database of the 12 species. We also demonstrated the practicability of the method in real fecal samples, achieved deep proteome coverage of human gut microbiome, and compared the function and taxonomy of gut microbiota at metagenomic level and metaproteomic level. Our study can tackle the current taxonomic annotation reliability problem in metagenomics-derived protein sequence database for metaproteomics. The unique dataset of metagenomic and the metaproteomic data of the 12 bacterial species is publicly available as a standard benchmarking sample for evaluating various analysis pipelines. The code of ConDiGA is open access at GitHub for the analysis of real microbiota samples.

bioinformatics↗

bioRxiv2023

Uncovering protein-protein interactions of the human sodium channel Nav1.7

The voltage-gated sodium channel Nav1.7 plays a crucial role in the initiation and propagation of pain signals. Our previous study has successfully identified the interacting proteins of mouse Nav1.7 (mNav1.7). In this study, we aimed to further elucidate the protein-protein interactions associated with human Nav1.7 (hNav1.7). Stable epitope (TAP)-tagged HEK293 cells expressing hNaV1.7 were utilized for the identification of hNav1.7-interacting proteins. The hNaV1.7-associated complexes were isolated through tandem affinity purification and further characterized by mass spectrometry. Bioinformatics analysis was carried out using the PANTHER classification system. Electrophysiological recording was performed to assess Nav1.7 current. Tap-tagged hNav1.7 was expressed effectively in HEK293 cells, exhibiting normal functional Nav1.7 currents. A total of 261 proteins were identified as interactors of hNav1.7, mainly located across the cell membrane and cytoplasm, and primarily involved in biological processes related to protein translation and expression. Comparison between human and mouse Nav1.7-interacting proteins revealed shared proteins (such as Eef1a1, Eef2, Tcp1, Cct2, Cct3, Cct5, Cct6a, and Cct7) as well as protein families (such as kinesin and Rab GTPases family). Knockdown of two of the shared interacting proteins, CCT5 and TMED10, resulted in reduced Nav1.7 current density. In conclusion, the protein interactions of hNaV1.7 were successfully mapped in the current work. These novel findings offer essential insights into the regulatory mechanisms that govern Nav1.7 function. Significance statementChronic pain affects approximately 20% of the worlds population and is a global major public health problem. Nav1.7 has been recognized as a promising target for novel analgesics. However, the drug development process for Nav1.7 is challenging. A thorough understanding of the regulatory mechanism of Nav1.7 would greatly assist in the development of its analgesic drugs. Our previous work successfully mapped the mNav1.7 protein interactions. In the current study, the interacting proteins of hNav1.7 were further defined. Our findings provide important implications for the development of Nav1.7-based analgesics for human use.

neuroscience↗

bioRxiv2023

Deletion of Pax1 scoliosis-associated regulatory elements leads to a female-biased tail abnormality

Adolescent idiopathic scoliosis (AIS), a sideways curvature of the spine, is sexually dimorphic, with increased incidence in females. A GWAS identified a female-specific AIS susceptibility locus near the PAX1 gene. Here, we used mouse enhancer assays, three mouse enhancer knockouts and subsequent phenotypic analyses to characterize this region. Using mouse enhancer assays, we characterized a sequence, PEC7, that overlaps the AIS-associated variant, and found it to be active in the tail tip and intervertebral disc. Removal of PEC7 or Xe1, a known sclerotome enhancer nearby, and deletion of both sequences led to a kinky phenotype only in the Xe1 and combined (Xe1+PEC7) knockouts, with only the latter showing a female sex dimorphic phenotype. Extensive phenotypic characterization of these mouse lines implicated several differentially expressed genes and estrogen signaling in the sex dimorphic bias. In summary, our work functionally characterizes an AIS-associated locus and dissects the mechanism for its sexual dimorphism.

genetics↗

bioRxiv2023

Doxorubicin inhibits phosphatidylserine decarboxylase and confers broad-spectrum antifungal activity

O_LIAs phospholipids of cell membranes, phosphatidylethanolamine (PE) and phosphatidylserine (PS) play crucial roles in glycerophospholipid metabolism. Broadly, some phospholipid biosynthesis enzymes serve as potential fungicide targets. Therefore, revealing the functions and mechanism of PE biosynthesis in plant pathogens would provide potential targets for crop disease control. C_LIO_LIWe performed analyses including phenotypic characterizations, lipidomics, enzyme activity, site-directed mutagenesis, and chemical inhibition assays to study the function of PS decarboxylase-encoding gene MoPSD2 in rice blast fungus Magnaporthe oryzae. C_LIO_LIThe Mopsd2 mutant was defective in development, lipid metabolism and plant infection. The PS level increased while PE decreased in Mopsd2, consistent with the enzyme activity. Furthermore, chemical doxorubicin inhibited the enzyme activity of MoPsd2 and showed antifungal activity against ten phytopathogenic fungi including M. oryzae and reduced disease severity of two crop diseases in the field. Three predicted doxorubicin-interacting residues are important for MoPsd2 functions. C_LIO_LIOur study demonstrates that MoPsd2 is involved in de novo PE biosynthesis and contributes to the development and plant infection of M. oryzae and that doxorubicin shows broad-spectrum antifungal activity as a fungicide candidate. The study also implicates that bacterium Streptomyces peucetius, which biosynthesizes doxorubicin, could be potentially used as an eco-friendly biocontrol agent. C_LI

pathology↗

bioRxiv2023

Δ246p53 is a new p53 isoform that responds to DNA damage and regulates tumour growth

p53 is with little doubt one of the most powerful genes in our genome, as it makes growth vs arrest, repair vs replacement, catabolism vs anabolism, life vs death decisions in the cell. An alteration or malfunction in p53 may lead to cancer, degeneration or premature ageing. So, it is not surprising that p53 is also one of the most complex and tightly regulated genes, encoding for at least 10 RNA variants and 12 widely accepted protein forms. Here we identify one new p53 protein isoform of approximately 18 kDa that we termed {Delta}246p53. {Delta}246p53 is translated from an alternative translation initiation site (TIS) in codon 246. TIS-246 is preceded by a strong Kozak sequence and appears conserved in vertebrates, from sea lamprey to humans. {Delta}246p53s origin and expression in cells were confirmed by frameshift and start codon mutations; five different antibodies against several epitopes from its N-terminus down to its C-terminus and one against the DNA binding domain region just upstream of its initiator methionine working as negative control; as well as siRNAs and an antisense oligo targeting TIS-246, which knocked-down {Delta}246p53 with little or no effect on full-length (FL) p53 protein levels. {Delta}246p53 was induced by DNA damage and triggered senescence and impaired tumour formation/growth in colony formation assays. Mechanistically, {Delta}246p53 interacted with FLp53, leading to a decrease in the expression of downstream target genes HDM2 and p21. Concurrently, it also interacted with {Delta}Np63, leading to p53-independent p63-mediated activation of p21, a known senescence activator. Our results unveil a new naturally occurring and tightly controlled factor with specific functions in p21 regulation and senescence. Future studies on {Delta}246p53 are likely to help us better understand and control the processes of tumour suppression and ageing.

cancer biology↗

bioRxiv2023

Strigolactones optimise plant water usage by modulating vessel formation

Wood formation is fundamental for the remarkable growth of plant bodies by continuously providing cells for long-distance transport of water and nutrients1-3. Vessel elements, the water transporting units within woody tissues, are generated from a cylindrical domain of cambium stem cells (CSCs) producing different vascular cell types in a bidirectional manner4-6. However, knowledge on the regulation of CSC activity is limited with unclear cell fate trajectories as the most obscure aspect in this context. Here, via revealing transcriptome signatures of CSCs and their derivatives with single cell resolution in Arabidopsis thaliana, we discover that the strigolactone (SL) signalling pathway modulates cell type composition in vascular tissues and thereby increases drought resistance. In particular, we find that SL signalling negatively regulates vessel element formation and thereby plant water usage. SL signalling is generally associated with differentiating vascular tissues but low in developing vessels and in CSCs implying a local role during fate decisions in CSC-derived cells. Highlighting the importance of vascular tissue composition for the overall plant water balance, alteration of vessel element formation has a direct impact on transpiration rates through leaf stomata. Our results demonstrate the importance of structural alignment of water transporting tissues to unstable water regimes and provide perspectives for a long-term modulation of drought resistance in plants.

plant biology↗

bioRxiv2023

Plant Arbovirus Mobilizes a Vector Salivary Protein to Initiate Plant Infection

Plant viruses typically rely on insect vectors for transmission between plants, with insect salivary proteins playing critical roles in this process. In this study, we demonstrate how Laodelphax striatellus salivary carbonic anhydrase (LssaCA) promotes rice stripe virus (RSV) infection in plants. We discovered that LssaCA directly binds to RSV nucleocapsid protein (NP) in insect salivary glands. This LssaCA-NP complex interacts with a rice thaumatin-like protein (OsTLP) that possess endo-{beta}-1,3-glucanase activity potentially degrading callose. Upon binding, the LssaCA-NP complex significantly enhances OsTLP enzymatic activity. We further clarify that both L. striatellus feeding and RSV infection induce callose deposition. The tripartite LssaCA-NP-OsTLP interaction enhances callose degradation, thereby facilitating RSV infection via its insect vector. This study provides new insights into complex virus-insect-plant tripartite interactions mediated by insect salivary proteins, with broad implications for numerous plant viruses transmitted by insect vectors.

plant biology↗

bioRxiv2023

Mutation nsp6 L232F associated with MERS-CoV zoonotic transmission confers higher viral replication in human respiratory tract cultures ex-vivo

Middle East respiratory syndrome coronavirus (MERS-CoV) causes zoonotic disease. Dromedary camels are the source of zoonotic infection. We identified a mutation of amino acid leucine to phenylalanine in the codon 232 position of the non-structural protein 6 (nsp6) (nsp6 L232F) that is repeatedly associated with zoonotic transmission. We generated a pair of isogenic recombinant MERS-CoV with nsp6 232L and 232F residues, respectively, and showed that the nsp6 L232F mutation confers higher replication competence in ex-vivo culture of human nasal and bronchial tissues and in lungs of mice experimentally infected in-vivo. Mechanistically, the nsp6 L232F mutation appeared to modulate autophagy and was associated with higher exocytic virus egress, while innate immune responses and zippering activity of the endoplasmic reticulum remained unaffected. Our study suggests that MERS-CoV nsp6 may contribute to viral adaptation to humans. This highlights the importance of continued surveillance of MERS-CoV in both camels and humans. ImportanceViral host adaptation plays an important role in zoonotic transmission of coronaviruses. For MERS-CoV that widely circulates in dromedary camels from Arabian Peninsula, camel-to human transmissions are ongoing, raising the possibility of human adaptive evolution for MERS-CoV. Here, we analysed MERS-CoV sequences and identified an amino acid mutation L232F in nsp6 to occur repeatedly in human MERS-CoV over the years since the first outbreak in 2012. We found the nsp6 L232F confers increase viral replication in-vitro, in ex-vivo upper human respiratory tract cultures and in mice, using a reverse genetics approach. Our results showed the nsp6 L232F may be advantageous for MERS-CoV to replicate in humans. This study highlighted a human adaptation of MERS-CoV and a need for continued surveillance of MERS-CoV to identify any further adaptations in humans, which may be relevant to the pandemic potential of MERS-CoV.

microbiology↗