Biology subjects

Mashruwala, A. A.

Publications and source records attributed to Mashruwala, A. A..

7 recordsLinked to original sources

bioRxiv2025

A transcription factor-sRNA-mediated double-negative feedback loop confers pathogen-specific control of quorum-sensing genes

The cell-to-cell communication process called quorum sensing enables bacteria to synchronize collective behaviors. Quorum sensing relies on the production, release, and detection of signaling molecules called autoinducers. In Vibrio cholerae, the VqmA transcription factor, following binding of the DPO autoinducer, activates expression of the gene encoding the VqmR small regulatory RNA. VqmR controls traits including biofilm formation. Here, we identify repressors of DPO-VqmA-VqmR signaling. We focus on one identified repressor, the LuxT transcription factor. We show that LuxT represses vqmR transcription. VqmR post-transcriptionally represses luxT translation. This arrangement forms a double-negative feedback loop between the two regulators. Reciprocal control hinges on the N-terminal 8 amino acids of LuxT. The nucleotide sequence encoding this LuxT region serves as the VqmR binding site in the luxT mRNA and the amino acids specified by this same N-terminal region are required for LuxT to bind the vqmR promoter. This same LuxT N-terminal region also expands the DNA motifs to which LuxT can bind. We show this regulatory circuit is unique to V. cholerae and closely related species and absent from other vibrios. We define the set of LuxT-controlled genes in V. cholerae and show that LuxT promotes biofilm formation, a key requirement for successful colonization of eukaryotic hosts. ImportanceBacterial quorum sensing enables control of collective behaviors. In Vibrio cholerae, the DPO-VqmA-VqmR quorum-sensing circuit governs key processes, including biofilm formation. Here, we identify a double-negative feedback loop between the transcription factor LuxT and the small RNA VqmR. This regulatory circuit depends on an eight amino acid N-terminal region that exists only in V. cholerae LuxT and LuxT from its close relatives. This short peptide sequence confers three distinct functions: It enables LuxT to repress vqmR, renders luxT mRNA susceptible to VqmR repression, and governs which DNA motifs LuxT can bind. Our findings reveal a pathogen-specific regulatory module that links small RNA targeting of mRNAs to transcription factor DNA binding specificity. The results show how evolution tailors bacterial regulatory circuits to adapt to different environments.

microbiology↗

bioRxiv2024

Quorum sensing orchestrates parallel cell death pathways in Vibrio cholerae via Type 6 secretion dependent and independent mechanisms

Quorum sensing (QS) is a cell-to-cell communication process that enables bacteria to coordinate group behaviors. In Vibrio cholerae colonies, a program of spatial-temporal cell death is among the QS-controlled traits. Cell death occurs in two phases, first along the colony rim, and subsequently, at the colony center. Both cell death phases are driven by the type VI secretion system (T6SS). Here, we show that HapR, the master QS regulator, does not control t6ss gene expression nor T6SS-mediated killing activity. Nonetheless, a {Delta}hapR strain displays no cell death at the colony rim. RNA-Seq analyses reveal that HapR activates expression of an operon containing four genes of unknown function, vca0646-0649. Epistasis and overexpression studies show that two of the genes, vca0646 and vca0647, are required to drive cell death in both a {Delta}hapR and a {Delta}hapR {Delta}t6ss strain. Thus, vca0646-0649 are regulated by HapR but act independently of the T6SS machinery to cause cell death, suggesting that a second, parallel pathway to cell death exists in V. cholerae. SignificanceCell death is a fundamental biological process. In mammals, cell death sculpts tissues during development, enables injury recovery, and regulates immunity. In bacteria, cell death mechanisms remain little explored. Recently, colonies formed by the pathogen Vibrio cholerae were demonstrated to undergo a spatio-temporal program of cell death. The program is controlled by quorum sensing (QS) and driven by the Type VI secretion system. Here, we discover QS-controlled genes, called vca0646-0649, that cause cell death in V. cholerae colonies independently of the Type VI secretion system. These findings indicate that a second cell death pathway exists in V. cholerae. The results expand our understanding of bacterial cell death mechanisms and provide insight into how cell death shapes bacterial community structure.

microbiology↗

bioRxiv2022

Quorum-sensing- and type VI secretion-mediated spatio-temporal cell death drives genetic diversity in Vibrio cholerae

Bacterial colonies composed of genetically identical individuals can diversify to yield variant cells with distinct genotypes. Variant outgrowth manifests as sectors. Here, we show that Type 6 secretion system (T6SS)-driven cell death in Vibrio cholerae colonies imposes a selective pressure for the emergence of variant strains that can evade T6SS-mediated killing. T6SS-mediated cell death occurs in two distinct spatio-temporal phases, and each phase is driven by a particular T6SS toxin. The first phase is regulated by quorum sensing and drives sectoring. The second phase does not require the T6SS-injection machinery. Variant V. cholerae strains isolated from colony sectors encode mutated quorum-sensing components that confer growth advantages by suppressing T6SS-killing activity while simultaneously boosting T6SS-killing defenses. Our findings show that the T6SS can eliminate sibling cells suggesting a role in intra-specific antagonism. We propose that quorum-sensing-controlled T6SS-driven killing promotes V. cholerae genetic diversity, including in natural habitats and during disease.

microbiology↗

bioRxiv2021

Bacterial LomR Induces the Vibriophage VP882 VqmA-Directed Quorum-Sensing Lysogeny-Lysis Transition

The bacterial cell-cell communication process called quorum sensing enables groups of bacteria to synchronously alter behavior in response to changes in cell population density. Quorum sensing relies on the production, release, accumulation, and detection of extracellular signal molecules called autoinducers. Here, we investigate a mechanism employed by a vibriophage to surveil host quorum sensing and tune its lysogeny-lysis decision to host cell density. The phage possesses a gene called vqmAPhage encoding a quorum-sensing receptor homologous to vibrio VqmA. Both VqmA receptors can detect the host bacteria-produced autoinducer called DPO. DPO-bound VqmAPhage launches the phage lysis process. We discover that the bacterial host produces an inducer of the VqmAPhage-directed quorum-sensing lysogeny-lysis transition. Production of the inducer appears to be widespread among bacteria. A screen of the Escherichia coli Keio collection for mutants impaired for inducer production revealed lomR, located in a prophage, and encoding a poorly understood protein. In the E. coli screening strain, lomR is interrupted by DNA encoding an insertion element. The 3 domain of this LomR protein is sufficient to induce VqmAPhage-directed lysis. Alanine-scanning mutagenesis showed that substitution at either of two key residues abrogates inducer activity. Full-length LomR is similar to the outer membrane porin OmpX in E. coli and Vibrio parahaemolyticus O3:K6, and OmpT in Vibrio cholerae C6706, and indeed, OmpX and OmpT can induce VqmAPhage-directed activity. Possibly, development of the LomR, OmpX, or OmpT proteins as tools to direct phage lysis of host cells could be used to control bacteria in medical or industrial settings. ABSTRACT IMPORTANCEBacteria communicate with chemical signal molecules using a process called quorum sensing. Quorum sensing allows bacteria to track their cell numbers and orchestrate collective behaviors. Recently, we discovered that a virus that infects and kills bacteria "eavesdrops" on its hosts quorum-sensing process. Specifically, the virus monitors host cell growth by detecting the accumulation of host quorum-sensing signal molecules. In response to the garnered quorum-sensing information, the virus kills the host bacterial cells when the bacterial population has reached a high cell density. This strategy presumably enhances transmission of viruses to new host cells. Here, we discover and characterize three closely-related bacterial host-produced proteins called LomR, OmpX, and OmpT that are capable of inducing the viral quorum-sensing-mediated killing program. Development of this class of inducer proteins as tools to drive "on demand" virus-mediated lysis of pathogenic host bacterial cells could be used to control bacteria in medical or industrial settings.

microbiology↗

bioRxiv2021

Secreted Proteases Control the Timing of Aggregative Community Formation in Vibrio cholerae

Bacteria orchestrate collective behaviors using the cell-cell communication process called quorum sensing (QS). QS relies on the synthesis, release, and group-wide detection of small molecules called autoinducers. In Vibrio cholerae, a multicellular community aggregation program occurs in liquid, during stationary phase, and in the high-cell-density QS state. Here, we demonstrate that this aggregation program consists of two subprograms. In one subprogram, which we call void formation, structures form that contain few cells but provide a scaffold within which cells can embed. The other subprogram relies on flagellar machinery and enables cells to enter voids. A genetic screen for factors contributing to void formation, coupled with companion molecular analyses, showed that four extracellular proteases, Vca0812, Vca0813, HapA, and PrtV control the onset timing of both void formation and aggregation, and moreover, proteolytic activity is required. These proteases, or their downstream products, can be shared between void-producing and non-void-forming cells and can elicit aggregation in a normally non-aggregating V. cholerae strain. Employing multiple proteases to control void formation and aggregation timing could provide a redundant and irreversible path to commitment to this community lifestyle. ImportanceBacteria can work as collectives to form multicellular communities. Vibrio cholerae, the bacterium that causes the disease cholera in humans, forms aggregated communities in liquid. Aggregate formation relies on a chemical communication process called quorum sensing. Here we show that, beyond overarching control by quorum sensing, there are two aggregation subprograms. One subprogram, which we call void formation, creates a scaffold within which cells can embed. The second subprogram, which allows bacteria to enter the scaffold, requires motility. We discovered that four extracellular proteases control the timing of both void formation and aggregation. We argue that, by using redundant proteases, V. cholerae ensures the reliable execution of this community formation process. These findings may provide insight into how V. cholerae successfully alternates between the marine environment and the human host: transitions that are central to the spread of the disease cholera.

microbiology↗

bioRxiv2020

The Vibrio cholerae quorum-sensing protein VqmA integrates cell density, environmental, and host-derived cues into the control of virulence

Scientific AbstractQuorum sensing is a chemical communication process in which bacteria use the production, release, and detection of signal molecules called autoinducers to orchestrate collective behaviors. The human pathogen Vibrio cholerae requires quorum sensing to infect the small intestine. There, V. cholerae encounters the absence of oxygen and the presence of bile. We show that these two stimuli differentially affect quorum sensing function and, in turn, V. cholerae pathogenicity. The quorum-sensing receptor-transcription factor called VqmA, that detects the autoinducer called DPO, also detects the lack of oxygen and the presence of bile. Detection occurs via DPO-, oxygen-, bile-, and redox-responsive disulfide bonds that alter VqmA DNA binding activity. We propose that VqmA serves as an information processing hub that integrates quorum- sensing information, redox status, the presence or absence of oxygen, and host cues. In response to the information acquired through this mechanism, V. cholerae appropriately modulates its virulence output. Lay AbstractQuorum sensing (QS) is a process of chemical communication bacteria use to orchestrate collective behaviors. QS communication relies on chemical signal molecules called autoinducers. QS regulates virulence in Vibrio cholerae, the causative agent of the disease cholera. Transit into the human small intestine, the site of cholera infection, exposes V. cholerae to the host environment. In this study, we show that the combination of two stimuli encountered in the small intestine, the absence of oxygen and the presence of host-produced bile, impinge on V. cholerae QS function and, in turn, pathogenicity. We suggest that possessing a QS system that is responsive to multiple environmental, host, and cell density cues enables V. cholerae to fine-tune its virulence capacity in the human intestine.

microbiology↗

bioRxiv2019

Drug-like Fragments Inhibit agr-Mediated Virulence Expression in Staphylococcus aureus

In response to the increasingly problematic emergence of antibiotic resistance, novel strategies for combating pathogenic bacteria are being investigated. Targeting the agr quorum sensing system, which regulates expression of virulence in Staphylococcus aureus, is one potentially useful approach for combating drug-resistant pathogens that has not yet been fully explored. A previously published study of a fragment screen resulted in the identification of five compound fragments that interact with the DNA-binding domain of the response regulator AgrA from S. aureus. We have analyzed the ability of these compounds to affect agr-mediated virulence gene expression in S. aureus cells. Three of the compounds demonstrated the ability to reduce agr-driven transcription of at the P2 and P3 promoters of the agr operon and increase biofilm formation, and two of these compounds also showed the ability to reduce levels of secreted toxins. The finding that the compounds tested were able to reduce agr activity suggests that they could be useful tools for probing the effects of agr inhibition. Furthermore, the characteristics of compound fragments make them good starting materials for the development of compound libraries to iteratively improve the inhibitors.

microbiology↗