Discipline: Biological Sciences
Christopher Rhoades - Michigan State University
Co-Author(s): Geoffrey Severin and Christopher Waters, Michigan State University, East Lansing, MI Christopher Waters, Michigan State University, East Lansing, Michigan
Vibrio cholerae, a pathogenic Gram-negative bacterium responsible for the disease cholera, utilizes a complex regulatory network to regulate virulence factor expression and pathogenicity. The latest V. cholerae biotype, El Tor, has supplanted Classical V. cholerae strains notably due to the acquisition of two novel gene islands, VSP-1 and VSP-2. These novel El Tor gene islands encode additional proteins necessary for biofilm formation and chemotaxis, of which both have been linked to the increased survivability of the bacterium within the gut of a human host. While the majority of genes in these islands remain to be characterized, the gene VC1079 encodes for a dinucleotide cyclase, DncV, whose predominant product is the hybrid cyclic nucleotide, cyclic-GMP-AMP (cGAMP). cGAMP is the newest described bacterial second messenger whose function has just begun to be explored. However, it has been shown that increased intracellular concentrations of cGAMP down regulate the chemotaxis genes in El Tor. Within a host, reduced chemotaxis is critical to El Tor’s pathogenicity but the molecular mechanisms involved in this cGAMP-mediated repression are unknown. We will determine this molecular pathway by inducing strong selective pressures for the evolution of chemotaxis mutants that resist cGAMP induced chemotactic inhibition. We have recently finished designing a high-throughput two-plasmid system that will more efficiently allow for identifying chemotactic mutants that have spontaneously mutated their genome, as opposed to a false positive from a plasmid derived mutation. Whole genome sequencing of these mutants will reveal genes involved in cGAMP mediated chemotactic repression. Chemotactic regulation is vital to V. cholerae pathogenicity and knowledge gained from this research will provide clues to understanding the enhanced pathogenic capacity of the El Tor biotype and cGAMP’s role in virulence.
Funder Acknowledgement(s): Funding for this research was obtained in part from Hunt for a Cure, awarded to Christopher Waters, Associate Professor for the Microbiology and Molecular Genetics Department,Michigan State University, East Lansing, Michigan.
Faculty Advisor: Christopher Waters, firstname.lastname@example.org
Role: In this process, I helped develop the screening system used to identify chemotactic mutants that may be resistant to cGAMP. I conducted all motility assays, screens, and quantified data and pictures used in this project. Images taken of the motility assays was also done by me, as well as culturing, growing, and plating bacteria used in this project.