The long-term objective of our research is to define signals that allow bacteria to communicate with a host and to identify the pathways by which they respond to the host environment. The symbiotic association between the Hawaiian squid, Euprymna scolopes, and its bacterial partner, Vibrio fischeri, provides a simple, elegant model system for studying bacteria-animal interactions. Only V. fischeri colonizes the squid. The evidence to date suggests that V. fischeri actively participates in achieving the observed specificity of the association. The factors that dictate this specificity, however, are not yet understood. We have identified a cluster of genes (syp, symbiosis polysaccharide locus) that is required for V. fischeri to initiate symbiosis, and an unlinked sensor kinase regulator, rscS, that controls syp transcription. Four additional regulators are proposed or known to also control syp transcription, including a ?54-dependent response regulator, SypG, and 2 additional 2-component regulators. Thus, syp is controlled by at least 3 proteins that are predicted to sense and respond to the environment. Multi-copy expression of a particular allele (rscS*) causes V. fischeri to express syp-dependent novel phenotypes consistent with altered cell-cell interactions: wrinkled colonies on solid complex media and pellicle formation in liquid minimal medium. We propose to elucidate the major regulatory mechanisms controlling syp transcription and identify additional genes associated with syp-dependent phenotypes. We will identify the polysaccharide produced by the syp locus and identify any differences in cell surface properties. Finally, we will examine in more detail the nature of the symbiosis defect of syp mutants and explore possible explanations to account for it. The experiments proposed here will expand our understanding of how colonization is initiated and how signal exchange occurs between a prokaryote and a eukaryote during the establishment of a long-term association. The relevance to public health lies in the potential of this model system to reveal novel mechanisms by which bacteria interact with an animal host. Such information could potentially allow the design of new antimicrobial agents. Our research organism is closely related to bacteria that cause gastroenteritis in humans, including the emerging pathogens, V. parahaemolyticus and V. vulnificus. Studying this model may also lead to approaches that prevent, reduce or treat such infections. ? ? ?
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