Stress activation of Bacillus subtilis sigma B can occur in the absence of the sigma B negative regulator RsbX

Abstract

Environmental stress activates B , the general stress response factor of Bacillus subtilis, by a pathway that is negatively controlled by the RsbX protein. To determine whether stress activation of B occurs by a direct effect of stress on RsbX, we constructed B. subtilis strains which synthesized various amounts of RsbX or lacked RsbX entirely and subjected these strains to ethanol stress. Based on the induction of a B -dependent promoter, stress activation of B can occur in the absence of RsbX. Higher levels of RsbX failed to detectably influence stress induction, but reduced levels of RsbX resulted in greater and longer-lived B activation. The data suggest that RsbX is not a direct participant in the B stress induction process but rather serves as a device to limit the magnitude of the stress response. B, the general stress response factor of Bacillus subtilis (20), is activated by a drop in intracellular ATP levels (1,22) or by any of a number of diverse environmental insults (e.g., heat shock, ethanol treatment, or salt shocks) (5, 7,22,23).B is held inactive by a binding protein (RsbW) which can form a complex with either B or an alternative target (RsbV) (6, 8, 9). RsbW is also a protein kinase which can phosphorylate RsbV and convert it into a form (RsbV-P) that no longer binds RsbW (8). The relatively high intracellular ATP levels that occur during growth favor the phosphorylation of RsbV, the formation of RsbW-B complexes, and the inhibition of B -dependent transcription (1,22). Upon entry into stationary phase, a drop in intracellular ATP levels leads to ineffective phosphorylation of RsbV, the formation of RsbV-RsbW complexes, and the release of B (1,22). Environmental stress activation of B occurs irrespective of intracellular ATP levels and the activity of the RsbW kinase (22). Instead, it involves the reactivation of RsbV-P by an RsbV-P-specific phosphatase (21). The existence of such a phosphatase was anticipated following the discovery that a phosphatase (SpoIIE) is involved in the reactivation of the phosphorylated form of the RsbV homolog (SpoIIAA) in the F system (2, 10). RsbV-P phosphatases participate in both stress-induced and stationaryphase activation of B (21). Phosphatase activity is essential for stress activation of B ; however, it merely enhances the stationary-phase response (21). Both the stationary-phase-and stress-induced dephosphorylation reactions require one or more of the products of the B regulators RsbR, RsbS, and RsbT (13,21,23). The RsbU protein is also needed for stressdependent, but not stationary-phase-dependent, RsbV-P dephosphorylation (21). Recent experiments have shown that RsbU can dephosphorylate RsbV-P in vitro and that this activity is enhanced by RsbT (15,24). It is likely that RsbU is a stress-activated phosphatase with RsbT providing the activation signal (24). The stationary-phase-specific phosphatase is unknown.An additional protein (RsbX) is a negative regulator of RsbU-dependent RsbV-P dephosphorylation. B. subtilis strains with null mutations in RsbX h...

In Vivo Phosphorylation of Partner Switching Regulators Correlates with Stress Transmission in the Environmental Signaling Pathway ofBacillus subtilis

“…For each stress, the amount of this modified form then decreased, reverting to a new steady-state level that was somewhat higher than that found in prestress cells. This decrease was presumably due to the action of the RsbX feedback phosphatase (24,27,30). Because the peak level of RsbS-P was detected at the first time point, the amount produced may have been underestimated.…”

“…This is particularly the case with RsbS-P, which achieves its highest measured level by the first time point, taken 1 min after the stress. Moreover, RsbS-P is labile to the action of the RsbX feedback phosphatase (9,27,30). RsbX expression is induced by stress in a B -dependent manner (15), and RsbX activity may also increase in response to stress (24).…”

“…RsbU then communicates the stress signal to the downstream partner switch by dephosphorylating RsbV-P, resulting in the activation of B (28,30). This model further holds that the environmental signal is damped by the RsbX feedback phosphatase (24,27,30), whose expression is under B control (15). The biochemical and genetic evidence supporting this model includes a consideration of the phenotypes caused by alteration of the residues upon which RsbS and RsbRA are phosphorylated (2,12,16,18,30).…”

Alternative transcription factor sigmaSB of Staphylococcus aureus: characterization and role in transcription of the global regulatory locus sar

“…B , identified in sporulating Bacillus subtilis, is the first member of the alternate factors to be reported in prokaryotes (20,21). Much has been learnt recently about the regulation of expression of the sigB operon and the modulation of the function of the SigB ( B ) protein in response to changing stress (entry into stationary growth phase, nutrient depletion, and presence of uncouplers of oxidative phosphorylation) and environmental stress (heat shock and osmotic shock) in B. subtilis (2,6,8,22,36,37,41).The emergence of multiple-drug-resistant, highly pathogenic strains of Staphylococcus aureus poses a serious threat for human and animal health management of infected subjects. S. aureus causes a number of infections ranging from superficial skin abscesses to life-threatening systemic diseases (e.g., toxic shock syndrome and osteomyelitis) (17,25,38).…”

The Bacillus subtilis GTP Binding Protein Obg and Regulators of the ς B Stress Response Transcription Factor Cofractionate with Ribosomes

“…RsbR is thought to mediate the RsbT-RsbS interaction, but its exact role in this process is not clear (1,10). The stress induction of B is limited by RsbX, which can dephosphorylate RsbS-P and reestablish the RsbS-dependent inhibition of RsbT (29,36,42).…”