Differential degradation of Escherichia coli sigma32 and Bradyrhizobium japonicum RpoH factors by the FtsH protease.

The Escherichia coli heat shock sigma factor sigma32 (RpoH) is rapidly degraded under non-stress conditions. The integrity of the DnaK chaperone machinery and the ATP-dependent FtsH protease are required for sigma32 proteolysis in vivo. Bradyrhizobium japonicum expresses three sigma32-type transcription factors, RpoH1, RpoH2, and RpoH3, which are functional in E. coli. We compared the stability of these sigma factors with E. coli sigma32 stability. In E. coli C600 (wild-type), the half-lives of sigma32, RpoH1, RpoH2 and RpoH3 were 30 s, 7 min, 4 min and 4 min, respectively. The first three proteins were stabilized in ftsH mutant backgrounds, indicating that they are degraded by FtsH in the wild-type. Proteolysis of RpoH3 was FtsH-independent because this sigma factor was not stabilized in ftsH mutants. Interestingly, in a purified in vitro system, all four RpoH proteins were degraded by FtsH, indicating that in vivo protein degradation depends on additional cellular factors. Rationally designed point mutations of sigma32 and RpoH1 suggested that the highly conserved RpoH box does not play a major role in conferring stability to RpoH factors. Presumably, several regions distributed along the primary sequence of the sigma factor are important for FtsH-mediated proteolysis. Finally, we provide evidence that proteolysis of RpoH factors in vivo depends on the DnaK machinery, irrespective of the protease involved.