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Use of a reporter gene to follow high-pressure signal transduction in the deep-sea bacterium Photobacterium sp. strain SS9.

Center for Marine Biomedicine and Biotechnology, Scripps Institution of Oceanography, University of California, San Diego, La Jolla 92093-0202.

ABSTRACT

Photobacterium sp. strain SS9 is a deep-sea bacterium which modulates the abundances of several outer membrane proteins as a function of hydrostatic pressure. These proteins include the product of the previously cloned ompH gene (D. H. Bartlett, M. Wright, A. A. Yayanos, and M. Silverman. Nature (London) 342:572-574, 1989). Subsequent to conjugal plasmid delivery it was possible to cross an ompH::lacZ transcriptional fusion into the genome of SS9, replacing the wild-type ompH gene, generating strain EC10. EC10 is not impaired in growth at high pressure, indicating that under the growth conditions employed, OmpH is not required for baroadaptation. beta-Galactosidase production in EC10 is induced by high pressure to approximately the same extent that OmpH production is in the parental strain, SS9. Therefore, OmpH abundance appears to be primarily regulated at the transcriptional level. EC10 was used for the isolation of ompH regulatory mutants. Derivatives of EC10 which produce reduced levels of beta-galactosidase at both low and high pressure and which appeared to possess mutations outside the ompH::lacZ locus were obtained. All of these regulatory mutants displayed alterations in the high-pressure repression of a second outer membrane protein, designated OmpL, and two of the mutants were also deficient in the high-pressure induction of a third outer membrane protein, designated OmpI. The most dramatic phenotype was present in mutant EC1002, whose growth was extremely barosensitive. EC1002 is the first pressure-sensitive mutant ever isolated. Prolonged incubation of EC1002 at high pressure led to the accumulation of cells with wild-type growth characteristics at high pressure. These cells are suggested to possess suppressor mutations, as they remain deficient in beta-galactosidase production and maintain their high-pressure-adapted phenotype for many generations in the absence of high-pressure selection.

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