Biology subjects

Ma, L.

Publications and source records attributed to Ma, L..

At least 199 records · Page 11Linked 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

Anthracyclines inhibit SARS-CoV-2 infection

Vaccines and drugs are two effective medical interventions to mitigate SARS-CoV-2 infection. Three SARS-CoV-2 inhibitors, remdesivir, paxlovid, and molnupiravir, have been approved for treating COVID-19 patients, but more are needed, because each drug has its limitation of usage and SARS-CoV-2 constantly develops drug resistance mutations. In addition, SARS-CoV-2 drugs have the potential to be repurposed to inhibit new human coronaviruses, thus help to prepare for future coronavirus outbreaks. We have screened a library of microbial metabolites to discover new SARS-CoV-2 inhibitors. To facilitate this screening effort, we generated a recombinant SARS-CoV-2 Delta variant carrying the nano luciferase as a reporter for measuring viral infection. Six compounds were found to inhibit SARS-CoV-2 at the half maximal inhibitory concentration (IC50) below 1 M, including the anthracycline drug aclarubicin that markedly reduced viral RNA-dependent RNA polymerase (RdRp)-mediated gene expression, whereas other anthracyclines inhibited SARS-CoV-2 by activating the expression of interferon and antiviral genes. As the most commonly prescribed anti-cancer drugs, anthracyclines hold the promise of becoming new SARS-CoV-2 inhibitors. IMPORTANCEMicrobial metabolites are a rich source of bioactive molecules. The best examples are antibiotics and immunosuppressants that have transformed the practice of modern medicine and saved millions of lives. Recently, some microbial metabolites were reported to have antiviral activity, including the inhibition of Zika virus and Ebola virus. In this study, we discovered several microbial metabolites that effectively inhibit SARS-CoV-2 infection, including anthracyclines that have also been shown to inhibit other viruses including Ebola virus through enhancing interferon responses, which indicates potentially broad antiviral properties of these microbial metabolites and can lead to the discovery of pan-antiviral molecules.

microbiology↗

bioRxiv2023

Two RNA-binding proteins mediate the sorting of miR223 from mitochondria into exosomes

Fusion of multivesicular bodies (MVBs) with the plasma membrane results in the secretion of intraluminal vesicles (ILVs), or exosomes. The sorting of one exosomal cargo RNA, miR223, is facilitated by the RNA-binding protein, YBX1 (Shurtleff et al., 2016). We found that miR223 specifically binds a "cold shock" domain (CSD) of YBX1 through a 5 proximal sequence motif UCAGU that may represent a binding site or structural feature required for sorting. Prior to sorting into exosomes, most of the cytoplasmic miR223 resides in mitochondria. An RNA-binding protein localized to the mitochondrial matrix, YBAP1, appears to serve as a negative regulator of miR223 enrichment into exosomes. miR223 levels decreased in the mitochondria and increased in exosomes after loss of YBAP1. We observed YBX1 shuttle between mitochondria and endosomes in live cells. YBX1 also partitions into P body granules in the cytoplasm (Liu et al., 2021). We propose a model in which miR223 and likely other miRNAs are stored in mitochondria and are then mobilized by YBX1 to cytoplasmic phase condensate granules for capture into invaginations in the endosome that give rise to exosomes.

cell biology↗

bioRxiv2022

Alternative Complement Pathway Regulates Thermogenic Fat Function via Adipsin and Adipocyte-derived C3aR1 in a Sex-dependent Fashion

Thermogenesis in beige/brown adipose tissues can be leveraged to combat metabolic disorders such as type 2 diabetes and obesity. The complement system plays pleiotropic roles in metabolic homeostasis and organismal energy balance with canonical effects on immune cells and non-canonical effects on non-immune cells. The adipsin/C3a/C3aR1 pathway stimulates insulin secretion and sustains pancreatic beta cell mass. However, its role in adipose thermogenesis has not been defined. Here, we show that Adipsin knockout mice exhibit increased energy expenditure and white adipose tissue (WAT) browning. C3a, a downstream product of adipsin, is generated from complement component 3 and decreases Ucp1 expression in subcutaneous adipocytes. In addition, adipocyte-specific C3aR1 knockout male mice show enhanced WAT thermogenesis and increased respiration. In stark contrast, adipocyte-specific C3aR1 knockout female mice display decreased brown fat thermogenesis and are cold intolerant. Female mice express lower levels of Adipsin in thermogenic adipocytes and adipose tissues than males. C3aR1 is also lower in female subcutaneous adipose tissue than males. Collectively, these results reveal sexual dimorphism in the adipsin/C3a/C3aR1 axis in regulating adipose thermogenesis. Our findings establish a newly discovered role of the alternative complement pathway in adipose thermogenesis and highlight sex-specific considerations in potential therapeutic targets for metabolic diseases.

cell biology↗

bioRxiv2022

Social tolerance as strategy to resolve group conflicts: an experimental study on tug-of-war competition of burying beetles

In animal groups with dominance hierarchies, there often occurs a tug-of-war competition over resources and reproduction between dominants and subordinates, because neither is able to fully control the other. Consequently, individuals may mitigate within-group conflict, either by fighting others or by signalling their willingness to tolerate others. Nevertheless, how such a tolerance interaction evolves remains unclear. Here, we addressed this knowledge gap and tested the tug-of-war competition hypothesis, by investigating whether subordinates pay to stay in the group by helping dominants (pay-to-stay), and whether dominants pay costs by living with subordinates in the group (pay-from-staying). We used the burying beetles, Nicrophorus vespilloides, which compete with intra- and inter-specifics for valuable carcasses that are needed for reproduction. Multiple conspecifics can reproduce together through communal breeding, thereby enhancing benefits in terms of reproduction and resource defence against competitors. In communal associations, larger individuals are often dominant in carcass use and reproduction, whereas subordinates have restricted access to the carcass. Our findings show that cooperative subordinates paid costs by helping dominant breeders in carcass preparation in order to be tolerated (i.e. increased access towards the carcass) by dominant breeders, but subordinates did not increase their reproductive success by helping. Such tolerance was eliminated by a high interspecific competition with blowfly maggots. Our results also show that dominant males, but not dominant females, benefitted more from the presence of subordinates, partly due to a sex difference in the compensation strategy of dominants. Overall, our study demonstrates that a social tolerance occurring in situations with a tug-of-war competition could be a common strategy to resolve conflicts in animal societies.

ecology↗

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

Effect of rectified gap junctional electrical coupling and spatial distribution of biologically engineered pacemaking cells on ventricular excitation

AimBiologically engineered pacemaker, or bio-pacemaker, is a promising replacement for electronic pacemakers for treating cardiac dysfunction. Previous animal experimental studies, however, have not been able to accurately demonstrate the stability and efficiency of the bio-pacemaker yet. This study aimed to elucidate the underlying factors that affect bio-pacemakers performance and to discover possible optimising solutions to enable the potential use of bio-pacemaker therapy. Methods and resultsThe human ventricular myocytes model in this study followed the ten Tussuchers model in 2006, and the bio-pacemaker single cell model was modified based on it as what has been expatiated in our previous work. In tissue model, two factors were primarily evaluated for their effects on bio-pacemakers to pace and drive surrounding cardiac tissue: gap junction between bio-pacemaker cells (PMs) and adjacent ventricular myocytes (VMs) and the spatial distribution of bio-pacemakers. A suppressed gap junctional electrical coupling between and heterotypic gap junctions were simulated and a combination of them led to the best performance of the bio-pacemaker. Then, the pacemaking behaviours of three kinds of idealised PM-VM slices were simulated, in which an electrically isolated distribution of bio-pacemaker showed optimal drive capacities. Finally, a real human ventricular slice model was used to verified the conclusions in idealized tissues. ConclusionThis study develops a theory that weak-rectified electrical coupling and electrically isolated distribution can enhance the pacemaking efficiency of bio-pacemakers, which lays the groundwork for future research into therapeutic applications of bio-pacemakers. Author summaryBiologically engineered pacemakers are expected to be a substitute for electronic pacemakers because of their physiological superiority, but how to transform them for practical application remains challenging. In this paper, we presented a theoretical perspective on optimising biological pacemaking capability based on a computational simulation approach. By manipulating the gap junctional electrical coupling among bio-pacemaking cells and between the pacemaker and their surrounding cells, and controlling spatial distribution of bio-pacemaker, we demonstrated that an enhanced capacity of a bio-pacemaker can be achieved. The results of this study may provide a theoretical basis for the further clinical development of bio-pacemakers.

biophysics↗

bioRxiv2022

Widely targeted metabolomic analysis reveals differences in volatile metabolites among four Angelica species

Angelica L. has attracted global interest for its traditional medicinal uses and commercial values. However, few studies have focused on the metabolomic differences among the Angelica species. In this study, employing the widely targeted metabolomics based on gas chromatography-tandem mass spectrometry, the metabolomes of four Angelica species were analyzed (Angelica sinensis (Oliv.) Diels (A. sinensis), Angelica biserrata (R.H.Shan & Yuan) C.Q.Yuan & R.H.Shan (A. biserrata), Angelica dahurica (Hoffm.) Benth. & Hook.f. ex Franch. & Sav. (A. dahurica), Angelica keiskei Koidz. (A. keiskei)). A total of 698 volatile metabolites were identified and classified into fifteen different categories. The metabolomic analysis indicated that 7-hydroxycoumarin and Z-ligustilide were accumulated at significantly higher levels in A. sinensis, whereas the opposite pattern was observed for bornyl acetate. In addition, a high correspondence between the dendrogram of metabolite contents and phylogenetic positions was detected in the four species. This study provides a biochemical map for the exploitation, application and development of the Angelica species as medicinal plants or health-related dietary supplements.

plant biology↗

bioRxiv2022

Genome-wide Identification and Expression Profile Analysis of Laccase Family Genes in the Hypsizygus marmoreus

Laccase exists widely in plants and fungi. It is a copper-containing polyphenol oxidase that can degrade lignin, oxidate, and phenolic substances, inhibit heterophytes, promote fruiting body formation, and improve the quality of mushrooms. In this study, 18 laccase genes were identified from the whole genome of a white strain (HM62) of Hypsizygus marmoreus, and the mapping, structure, and evolution of laccase genes were analyzed at the whole genome level, while the spatiotemporal expression was evaluated at different developmental stages. The laccase genes mainly distributed on chromosomes 1, 2, 3, 4, 6, 9, and 10, and 9 genes were clustered linearly on chromosome 6, indicating gene doubling. Phylogenetic tree analysis showed that the laccase gene family was divided into three subfamilies. The spatiotemporal expression analysis of the laccase gene family showed that HmLac09 and HmLac10 were highly expressed in different periods and might be involved in lignin degradation and fruit body formation, respectively. The expression levels of HmLac02, HmLac05, HmLac08, and HmLac17 genes in gray or gray and white heterozygous strains were higher than those in white strains, which might be related to the difference in lignin decomposition in gray strains, and one of the factors leading to different growth rates. The present study investigated the characterization of the H. marmoreus laccase gene family, extending our understanding of laccase mediated fruiting body development and growth rate mechanisms in this fungi.

microbiology↗

bioRxiv2022

Characterization of N distribution in different organs of winter wheat using UAV-based remote sensing

Although unmanned aerial vehicle (UAV) remote sensing is widely used for high-throughput crop monitoring, few attempts have been made to assess nitrogen content (NC) at the organ level and its impact on nitrogen use efficiency (NUE). Also, little is known about the performance of UAV-based image texture features in crop nitrogen and NUE monitoring. In this study, eight flying missions were carried out throughout different stages of winter wheat (from the jointing stage to the stage 25 days after flowering) to acquire multispectral images. Forty-three multispectral vegetation indices (VIs) and forty texture features (TFs) were calculated from images and fed into the partial least squares regression (PLSR) and random forest (RF) regression models for predicting nitrogen-related indicators. Our main purposes were to (1) evaluate the potential of UAV-based images to predict NC in different organs of winter wheat and nitrogen agronomic efficiency (NAE); (2) compare the performances of VIs, TFs and the combination of them for nitrogen monitoring. The results showed that the correlation between different features (VIs and TFs) and NC in different organs varied between the vegetative and reproductive phases. Most of VIs were found to be positively correlated with NC, while most of the TFs were negatively correlated with NC. PLSR latent variables extracted from VIs and TFs explained 80% of the variations in NAE. However, no significant differences were found between VIs and TFs in their performance in predicting NC in different organs. This study demonstrated the promise of applying UAV-based imaging to estimate NC and NAE in different organs of winter wheat.

bioengineering↗

bioRxiv2022

USP10 strikes down Wnt/β-catenin signaling by dual-wielding deubiquitinase activity and phase transition potential

Wnt/{beta}-catenin signaling is a conserved pathway crucially governing development, homeostasis and oncogenesis. Discovery of novel regulators holds great values in both basic and translational research. Through screening, we identified a deubiquitinase (DUB) USP10 as a novel and critical modulator of {beta}-catenin. Mechanistically, USP10 binds to key scaffold Axin1 via conserved motifs and stabilizes Axin1 through K48-linked deubiquitination, and surprisingly, tethers Axin1 and {beta}-catenin physically while promoting phase separation for {beta}-catenin suppression regardless of its enzymatic activity. Functionally, USP10 prominently regulates embryonic development and intestinal homeostasis by antagonizing {beta}-catenin via DUB activity. In colorectal cancer, USP10 substantially represses cancer growth mainly through physical binding compensation and phase separation promotion and correlates with Wnt/{beta}-catenin magnitude clinically. Collectively, we discovered USP10 functioning in multiple biological processes against {beta}-catenin and unearthed a novel enzyme-dependent and -independent "dual-regulating" mechanism by which USP10 utilizes parallelly and context-dependently. USP10 inhibitor was suggested in treating certain Wnt-related diseases.

cell biology↗

bioRxiv2022

A non-human primate model with Alzheimer's disease-like pathology induced by hippocampal overexpression of human tau

Alzheimers disease (AD) is one of the most burdening diseases of the century with no disease-modifying treatment yet. Non-human primates (NHPs) share genetic, anatomical and physiological similarities with humans, making them an ideal model for investigating the pathogenesis and therapeutics of AD. However, the applications of NHPs in AD research have been hindered by the paucity of spontaneous or induced monkey models for AD due to their long generation time, ethical considerations and technical challenges in making genetically modified monkeys. Here we developed an AD-like NHP model by overexpressing human tau in bilateral hippocampi of adult rhesus macaque monkeys. We evaluated the pathological features of these monkeys with immunostaining, cerebrospinal fluid (CSF) analysis, magnetic resonance imaging (MRI), positron emission tomography (PET) scan, and behavioral tests. We demonstrated that after hippocampal overexpression of human tau, the rhesus macaque monkeys displayed multiple pathological features of AD, including neurofibrillary tangle formation, neuronal loss, hippocampal atrophy, neuroinflammation, A{beta} clearance deficit, blood vessel damage and cognitive decline. This work establishes a human tau-induced AD-like NHP model that may facilitate mechanistic studies and therapeutic treatments for AD.

neuroscience↗

bioRxiv2022

Macaque Brainnetome Atlas with Fine-grained Parcellations and Reliable Connections

The rhesus macaque (Macaca mulatta) is a crucial experimental animal that shares many genetic, brain organizational, and behavioral characteristics with humans. A macaque brain atlas that identifies anatomically and functionally distinct regions is fundamental to biomedical and evolutionary research. However, even though connectivity information is vital for understanding brain functions, a connectivity-based whole-brain atlas of the macaque has not previously been made. In this study, we created a new whole-brain map, the Macaque Brainnetome Atlas (MacBNA), based on the anatomical connectivity profiles provided by high angular and spatial resolution ex vivo diffusion MRI data. The new atlas consists of 248 cortical and 56 subcortical regions as well as their structural and functional connections. The parcellation and the diffusion-based tractography were comprehensively evaluated with multi-contrast MRI, invasive neuronal-tracing, and Nissl-stained images collected from a single subject and with open-access datasets from other cohorts. As a demonstrative application, the structural connectivity divergence between macaque and human brains was mapped using the Brainnetome atlases of those two species to uncover the genetic underpinnings of the evolutionary changes in brain structure. The resulting resource includes (1) the thoroughly delineated Macaque Brainnetome Atlas (MacBNA), (2) regional connectivity profiles, (3) the postmortem high resolution macaque diffusion and T2-weighted MRI dataset (Brainnetome-8), and (4) multi-contrast MRI, block-face, and section images collected from a single macaque. MacBNA can serve as a common reference frame for mapping multifaceted features across modalities and spatial scales and for integrative investigation and characterization of brain organization and function. Therefore, it will enrich the collaborative resource platform for nonhuman primates and facilitate translational and comparative neuroscience research.

neuroscience↗

bioRxiv2022

Domain Adaptive Neural Inference for Neurons, Microcircuits and Networks

Accurately inferring neural circuit connectivity from in vivo recordings is essential for understanding the computations that support behavior and cognition. However, current deep learning approaches are limited by incomplete observability and the lack of ground-truth labels in real experiments. Consequently, models are often trained on synthetic data, which leads to the well-known "model mismatch" problem when simulated dynamics diverge from true neural activity. To overcome these challenges, we present Deep Domain-Adaptive Matching (DeepDAM), a training framework that adaptively matches synthetic and in vivo data domains for neural connectivity inference. Specifically, DeepDAM fine-tunes deep neural networks on a combined dataset of synthetic simulations and unlabeled in vivo recordings, aligning the models feature representations with real neural dynamics to mitigate model mismatch. We demonstrate this approach in rodent hippocampal CA1 circuits as a proof-of-concept, achieving near-perfect connectivity inference performance (Matthews correlation coefficient [~]0.97-1.0) and substantially surpassing classical methods ([~]0.6-0.7). We further demonstrate robustness across multiple recording conditions within this hippocampal dataset. Additionally, to illustrate its broader applicability, we extend the framework to two distinct systems without altering the core methodology: a stomatogastric microcircuit in Cancer borealis (ex vivo) and single-neuron intracellular recordings in mouse, where DeepDAM significantly improves efficiency and accuracy over standard approaches. By effectively leveraging synthetic data for in vivo and ex vivo analysis, DeepDAM offers a generalizable strategy for overcoming model mismatch and represents a critical step towards data-driven reconstruction of functional neural circuits.

neuroscience↗

bioRxiv2022

Eradication of tumors with pre-existing antigenic heterogeneity by vaccine-mediated co-engagement of CAR T and endogenous T-cells

Chimeric Antigen Receptor (CAR) T-cell therapy can be effective in treating human cancer but loss of the antigen recognized by the CAR poses a major obstacle. Here, we report an approach for vaccine boosting CAR T-cells, which triggers engagement of the endogenous immune system to circumvent antigen-negative tumor escape. Vaccine-boosted CAR T-cells promoted dendritic cell (DC) recruitment to tumors, increased tumor antigen uptake by DCs, and elicited priming of endogenous anti-tumor T-cells (antigen spreading). This process was accompanied by a shift in toward oxidative phosphorylation in CAR T-cells and was critically dependent on CAR-T-derived IFN-{gamma}. Antigen spreading induced by vaccine-boosted CAR-T enabled a proportion of complete responses even when the initial tumor was 50% CAR-antigen-negative, and heterogenous tumor control was further enhanced by genetically amplifying CAR-T IFN-{gamma} expression. Thus, CAR T-cell-derived IFN-{gamma} plays a critical role in promoting antigen spreading, and vaccine boosting provides a clinically-translatable strategy to drive such responses against solid tumors.

immunology↗

bioRxiv2022

Development of SARS-CoV-2 mRNA vaccines encoding spike N-terminal and receptor binding domains

With the success of mRNA vaccines against coronavirus disease 2019 (COVID-19), strategies can now focus on improving vaccine potency, breadth, and stability. We present the design and preclinical evaluation of domain-based mRNA vaccines encoding the wild-type spike-protein receptor-binding (RBD) and/or N-terminal domains (NTD). An NTD-RBD linked candidate vaccine, mRNA-1283, showed improved antigen expression, antibody responses, and stability at refrigerated temperatures (2-8{degrees}C) compared with the clinically available mRNA-1273, which encodes the full-length spike protein. In mice administered mRNA-1283 as a primary series, booster, or variant-specific booster, similar or greater immune responses and protection from viral challenge were observed against wild-type, beta, delta, or omicron (BA. 1) compared with mRNA-1273 immunized mice, especially at lower vaccine dosages. These results support clinical assessment of mRNA-1283 (NCT05137236). One Sentence SummaryA domain-based mRNA vaccine, mRNA-1283, is immunogenic and protective against SARS-CoV-2 and emerging variants in mice.

immunology↗

bioRxiv2022

Alternative mRNA polyadenylation bridges mitochondrion-to-nucleus communication in Arabidopsis

Mitochondria produce signals besides energy and metabolites that influence plant growth and fitness. However, how mitochondrial signals are relayed to other cellular compartments is largely unknown. By applying poly(A)-site RNA-sequencing (PAS-seq) to wildtype Arabidopsis seedlings and a mutant in the histone demethylase JMJ30 treated with the mitochondrial electron transfer chain inhibitor antimycin A (AA), we identified a previously undefined mitochondrion-to-nucleus communication pathway by which mitochondrial functional state regulates co-transcriptionally alternative polyadenylation (APA) of nuclear mRNA. We observed a global shortening of 3' untranslated regions (UTRs) as a molecular signature of AA-activated mitochondrial retrograde response (MRR), which contributed in part to translational regulation of auxin response and cell wall biogenesis. JMJ30 regulated AA-induced 3' UTR shortening, resulting in more transcripts with shortened 3' UTRs upon AA treatment in a JMJ30 gain-of-function mutant and overexpression lines. We also report on the JMJ30-interacting protein CPSF30, a cleavage and polyadenylation specificity factor that recruits JMJ30 to modulate H3K27me3 status at its target loci. Our study illustrates how epigenetic modification and APA coordinate mitochondrion-to-nucleus communication to allow cells to rapidly respond to changes in mitochondrial functional state and shape plant growth and fitness. One-sentence summaryEpigenetic modification and APA coordinate mitochondrion-to-nucleus communication to allow cells to rapidly respond to changes in mitochondrial functional state and shape plant growth and fitness.

molecular biology↗

bioRxiv2022

From motor protein to toxin: Mutations in the zonula occludens toxin (Zot) of Vibrio cholerae phage CTXΦ suggest a loss of phage assembly function

Prophages, i.e. dormant viruses residing in bacterial cells, are not just passive passengers in the bacterial host. Several prophage-encoded genes have been shown to be contributors to bacterial virulence by mediating antimicrobial resistance or by providing toxins. Other prophage genes exhibit beneficial effects on the host by modulating e.g. motility or biofilm formation. In this study, we used an in vivo phage assembly assay and tested an extensive array of single point mutations or their combinations found in Zot, the zonula occludens toxin encoded by the Vibrio cholerae phage CTX{phi}. The assay makes use of the highly homologous Zot-like protein g1p of the filamentous Coliphage M13, a motor protein that mediates the trans-envelope assembly and secretion of filamentous phages. We also measured the in vitro ATP hydrolysis of purified proteins, and quantified virus production in V. cholerae mediated by Zot or the Zot-like protein of the two Vibrio phages CTX{phi} and VFJ{phi}. In addition, we investigated sequence variations of the Walker motifs in Vibrio species using bioinformatics method, and revealed the molecular basis of ATP binding using molecular docking and molecular dynamics simulation based on the structure predicted by AlphaFold2. Our data indicates that g1p proteins in Vibrio can easily accumulate deleterious mutations and likely lose the ability to efficiently hydrolyse ATP, while the CTX{phi} Zot was further exapted to now act as an auxiliary toxin during the infection by Vibrio cholerae.

microbiology↗