Regulation of IS492 transposition in Pseudoalteromonas atlantica
Higgins, Brian Patrick
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A variety of specialized DNA recombination systems create genetic diversity and regulate gene expression in prokaryotes and eukaryotes. The goal of this research was to characterize the regulation of an unusual specialized recombination system, reversible insertion and excision of the insertion sequence IS492. Transposition of IS492, which is mediated by the transposase MooV, controls the production of peripheral extracellular polysaccharide (EPS) in the Gram-negative marine organism Pseudoalteromonas atlantica. EPS production is necessary for the switch from a planktonic existence in ocean water columns to biofilm formation on solid surfaces and in sediment. This work demonstrates that the on/off phase variation of EPS is controlled by insertion and precise excision of IS492 from a single site within epsG, a glycosyl transferase gene that is essential for EPS production. In a novel application of real time polymerase chain reaction (qPCR) technology, the frequency of IS492 precise excision from -2 epsG was measured and found to be as high as 10 per cell per generation, which is unprecedented for a IS element. This high frequency precise excision is dependent on the chromosomal context of IS492 and on environmental factors. Analysis of the relevance of the chromosomal context for IS492 indicates that transcription through the element from an external promoter increases the level of transpose-encoding transcripts. Thus, a regulator epsG transcription would control excision of IS492; the best candidate for this regulator, based EPS phase variation and IS492 excision frequency assays under various growth conditions, is the primary building block of EPS, galactose.