Abstract
Natural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination. It represents one mode of horizontal gene transfer and contributes to the spread of traits like antibiotic resistance. In Vibrio cholerae, a type IVa pilus (T4aP) is thought to facilitate natural transformation by extending from the cell surface, binding to exogenous DNA, and retracting to thread this DNA through the outer membrane secretin, PilQ. Here, we use a functional tagged allele of VcPilQ purified from native V. cholerae cells to determine the cryoEM structure of the VcPilQ secretin in amphipol to ~2.7 Å. We use bioinformatics to examine the domain architecture and gene neighborhood of T4aP secretins in Proteobacteria in comparison with VcPilQ. This structure highlights differences in the architecture of the T4aP secretin from the type II and type III secretion system secretins. Based on our cryoEM structure, we design a series of mutants to reversibly regulate VcPilQ gate dynamics. These experiments support the idea of VcPilQ as a potential druggable target and provide insight into the channel that DNA likely traverses to promote the spread of antibiotic resistance via horizontal gene transfer by natural transformation.
Highlights
Natural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination
One mechanism of horizontal gene transfer is natural transformation, where a competent bacterium can take up DNA from its environment and maintain this exogenous genetic material, either as a plasmid or by integrating it into its genome by homologous recombination[2]
The genomes of V. cholerae, V. parahaemolyticus, and V. vulnificus all contain two T4aP systems: mannose-sensitive hemagglutinin (MSHA) pili and chitinregulated T4aP used for natural competence[21]; the latter of which allows for chitin-induced natural transformation[22,23,24,25,26]
Summary
Natural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination It represents one mode of horizontal gene transfer and contributes to the spread of traits like antibiotic resistance. Based on our cryoEM structure, we design a series of mutants to reversibly regulate VcPilQ gate dynamics These experiments support the idea of VcPilQ as a potential druggable target and provide insight into the channel that DNA likely traverses to promote the spread of antibiotic resistance via horizontal gene transfer by natural transformation. One mechanism of horizontal gene transfer is natural transformation, where a competent bacterium can take up DNA from its environment and maintain this exogenous genetic material, either as a plasmid or by integrating it into its genome by homologous recombination[2]. V. cholerae is the most genetically tractable and has emerged as a model system for studying natural transformation and bacterial pili
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