The cellular concentration of the sigma S subunit of RNA polymerase in Escherichia coli is controlled at the levels of transcription, translation, and protein stability.

Abstract

The second vegetative sigma factor crs (encoded by the rpoS gene) is the master regulator in a complex regulatory network that governs the expression of many stationary phase-induced and osmotically regulated genes in Escherichia coil Using a combination of gene-fusion technology and quantitative immunoblot, pulse-labeling, and immunoprecipitation analyses, we demonstrate here that rpoS/~r s expression is not only transcriptionally controlled, but is also extensively regulated at the levels of translation and protein stability. rpoS transcription is inversely correlated with growth rate and and is negatively controlled by cAMP-CRP. In complex medium rpoS transcription is stimulated during entry into stationary phase, whereas in minimal media, it is not significantly induced, rpoS translation is stimulated during transition into stationary phase as well as by an increase in medium osmolarity. A model involving mRNA secondary structure is suggested for this novel type of post-transcriptional growth phase-dependent and osmotic regulation. Furthermore, ~r s is a highly unstable protein in exponentially growing cells (with a half-life of 1.4 min), that is stabilized at the onset of starvation. When cells are grown in minimal glucose medium, translational induction and ~r s stabilization occur in a temporal order with the former being stimulated already in late exponential phase and the latter taking place at the onset of starvation. Although crs does not control its own transcription, it is apparently indirectly involved in a negative feedback control that operates on the post-transcriptional level. Our analysis also indicates that at least five different signals [cAMP, a growth rate-related signal (ppGpp?I, a cell density signal, an osmotic signal, and a starvation signal] are involved in the control of all these processes that regulate rpoS/~r s expression.

The LysR‐like regulator LeuO in Escherichia coli is involved in the translational regulation of rpoS by affecting the expression of the small regulatory DsrA‐RNA

“…leuO is located at 1.83 min of the current physical map of the E. coli chromosome (Berlyn et al, 1996), and the location of Tn10-8 at this position was confirmed by P1 (D, E and F). The fusions used were the transcriptional r poS742 ::lacZ fusion (A and D) and the translational r poS742 ::lacZ (B and E) and r poS379 ::lacZ (C and F) fusions (Lange and Hengge-Aronis, 1994;Muffler et al, 1996a). Optical densities (circles) and specific ␤-galactosidase activities (triangles) were determined throughout the growth cycle.…”

“…Samples of 40 g of total cellular protein were applied per lane. A polyclonal serum against S (Lange and Hengge-Aronis, 1994), a goat anti-rabbit IgG alkaline phosphatase conjugate (Sigma) and a chromogenic substrate (BCIP/NBT; Boehringer Mannheim) were used for the visualization of S bands.…”

“…Not only are rpoS transcription and translation differentially controlled, but S is also a very unstable protein whose rate of proteolysis is tightly regulated. S half-life varies between approximately 1.5 min and more than 40 min, depending on growth and specific stress conditions (Lange and Hengge-Aronis, 1994;Takayanagi et al, 1994;Muffler et al, 1996a;1997a). With the exception of entry into stationary phase after growth in complex medium, stress conditions hardly alter rpoS transcription, but various stress signals differentially affect either rpoS translation or S turnover or both.…”

“…by the addition of NaCl or sucrose (Muffler et al, 1996a), temperature downshift (Sledjeski et al, 1996) and during late exponential phase above a certain cell density, when cells are grown in minimal medium (Lange and Hengge-Aronis, 1994). S proteolysis is more or less strongly inhibited in response to starvation (Lange and Hengge-Aronis, 1994;Takayanagi et al, 1994), high osmolarity (Muffler et al, 1996a), heat shock (Muffler et al, 1997a) and, probably, shift to acid pH (Bearson et al, 1996). So far, osmotic upshift is the only known stress condition that at the same time stimulates rpoS translation and almost completely inhibits S degradation (Muffler et al, 1996a).…”

Heat‐shock and general stress response in Bacillus subtilis

“…This induced or adaptive protection depends on the action of s 32 , whose activity increases transiently after heat stress leading to the induction of the heatshock regulon. Furthermore, it is interesting to note that the expression of both sigma factors is controlled by similar mechanisms at translational as well as posttranslational level (for review see Yura et al, 1993;Lange and Hengge-Aronis, 1994).…”