Vibrio species, including the squid symbiont Vibrio fischeri, become competent to take up DNA under specific conditions. For example, V. fischeri becomes competent when grown in the presence of chitin oligosaccharides or upon overproduction of the competence regulatory factor TfoX. While little is known about the regulatory pathway(s) that control V. fischeri competence, this microbe encodes homologs of factors that control competence in the well-studied V. cholerae. To further develop V. fischeri as a genetically-tractable organism, we evaluated the roles of some of these competence homologs. Using TfoX-overproducing cells, we found that competence depends upon LitR, the homolog of V. cholerae master quorum sensing and competence regulator HapR, and on homologs of putative pili genes that in V. cholerae facilitate DNA uptake. Disruption of genes for negative regulators upstream of LitR, namely the LuxO protein and the sRNA Qrr1, resulted in increased transformation frequencies. Unlike LitR-controlled light production, however, competence did not vary with cell density under tfoX-overexpressing conditions. Analogous to V. cholerae, the requirement for LitR could be suppressed by loss of the Dns nuclease. We also found a role for the putative competence regulator CytR. Finally, we determined that transformation frequencies varied depending on the TfoX-encoding plasmid, and developed a new dual tfoX- and litR-overexpression construct that substantially increased the transformation frequency of a less genetically-tractable strain. By advancing the ease of genetic manipulation of V. fischeri, these findings will facilitate the rapid discovery of genes involved in physiologically-relevant processes, such as biofilm formation and host colonization.
Importance The ability of bacteria to take up DNA (competence) and incorporate foreign DNA into their genomes (transformation) permits them to rapidly evolve and gain new traits and/or acquire antibiotic resistances. It also facilitates laboratory-based investigations into mechanisms of specific phenotypes, such as those involved in host colonization. Vibrio fischeri has long been a model for symbiotic bacteria-host interactions as well as for other aspects of its physiology such as bioluminescence and biofilm formation. Competence of V. fischeri can be readily induced upon overexpression of the competence factor TfoX. Relatively little is known about the V. fischeri competence pathway, although homologs of factors known to be important in V. cholerae competence exist. By probing the importance of putative competence factors that control transformation of V. fischeri, this work deepens our understanding of the competence process and advances our ability to genetically manipulate this important model organism.
Physical control, transformation and damage in the First World War: War bodies
