Transcript analysis of the c-vac region and differential synthesis of the two regulatory gas vesicle proteins GvpD and GvpE in Halobacterium salinarium PHH4.

Abstract

Halobacterium salinarium PHH4 synthesizes gas vesicles in the stationary growth phase by the expression of 14 gyp genes arranged in two clusters. The chromosomal gvpACNO (c-gvpACNO) gene cluster (encoding the major structural gas vesicle protein GvpA and the minor structural protein GvpC was transcribed as three mRNA species starting at one promoter during the stationary phase of growth. The second gene cluster, c-gvpDEFGHIKLM), was transcribed during all stages of growth as a relatively unstable, single mRNA with a maximal length of 6 kb. In addition, a 1.7-kb c-gvpD transcript was synthesized during stationary growth starting at the same promotor as that of the cgvpDEFGHIJKLM mRNA. The expression of the first two genes located in this unit (c-gvpD and c-gvpE) was also monitored by Western blot (immunoblot) analyses using antisera raised against these proteins synthesized in Escherichia coli. While the cGvpD protein was present only during early exponential growth and disappeared during gas vesicle formation, the cGvpE protein was present during cGvpA and gas vesicle synthesis in the early stationary phase of growth. Previous data indicated that cGvpD is involved in repression of gas vesicle formation, whereas cGvpE is a transcriptional activator for the c-gvpA promoter. The appearance of both proteins during the growth cycle is in line with the functions of these proteins in gas vesicle synthesis. The mechanism of the differential translation of cGvpD and cGvpE from the c-gvpDEFGHIJKLM rnRNA still has to be elucidated, but antisense RNAs complementary to the 5' terminus as well as the 3' portion of the c-gvpD mRNA might be involved in this regulation. Such RNAs occurred during early stationary growth when the cGvpD protein level decreased and may possibly inhibit the translation of the c-gvpD mRNA.