Transposition of IS492 and Regulation of Expression of Extracellular Polysaccharide in Pseudoalteromonas atlantica

University Of Georgia Research Foundation Inc, Athens GA

Investigators

Abstract

The marine bacterium Pseudoalteromonas atlantica switches between an extracellular polysaccharide (eps)-producing and an eps non-producing state. The switch is mediated by a precise insertion and excision of the transposon I S492. The proposed research will characterize transposition of the insertion element and determine how reversible insertion of IS492 at the eps locus regulates extracellular polysaccharide (EPS) synthesis. The precise insertion of IS492 at its target site within the eps gene eliminates extracellular polysaccharide production and excision of the transposon from eps restores eps expression. The transposition process is novel in that it is responsive to growth conditions, such as cell density. Because of this, the regulation of transposition is likely to be novel. The research will first define the essential steps in transposition using PCR-based assays and genetic selections/screens for IS492 insertion/excision at the eps locus in P. atlantica and on a plasmid in E. coli (Balding-Perkins et al., J. Bacteriol. 181(16), 1999). As these steps are determined, the regulation of transposition and the impact on eps expression will be assayed by measuring the frequency of transposition of IS492 in response to various environmental signals. Primer extension assays for eps mRNA and genetic fusions for eps expression will be used to identify the regulatory link between IS492 transposition and EPS production. Studying the regulation of EPS production by marine bacteria is extremely important to our understanding of the establishment of many types of marine ecosystems. It has been shown that the EPS produced by marine bacteria, including P. atlantica, facilitates the development of microbial biofilms. P. atlantica is an excellent organism for both basic research and research on marine microbial communities because it is culturable and amenable to genetic and biochemical characterization, whereas many of the other marine bacteria are not. The proposed research aimed at understanding the regulation of transposition and the role of this transposition in extracellular polysaccharide synthesis, therefore, is essential to our understanding of the broader picture of marine microbial communities.