Dynamic interplay between antagonistic pathways controlling the sigma(32) level in Escherichia coli

The heat-shock response in Escherichia coli depends primarily on the transi ent increase in the cellular level of heat-shock sigma factor sigma(32) enc oded by the rpoH gene, which results from both enhanced synthesis and trans ient stabilization of normally unstable sigma(32). Heat-induced synthesis o f sigma(32) was previously shown to occur at the translation level by melti ng the mRNA secondary structure formed within the 5' coding sequence of rpo H including the translation initiation region. The subsequent decrease in t he sigma(32) level during the adaptation phase has been thought to involve both shutoff of synthesis (translation) and destabilization of sigma(32)-me diated by the DnaK-DnaJ chaperones, although direct evidence for translatio nal repression was lacking. We now show that the heat-induced synthesis of sigma(32) does not shut off at the translation level by using a reporter sy stem involving translational coupling. Furthermore, the apparent shutoff wa s not observed when the synthesis rate was determined by a very short pulse labeling (15 s). Examination of sigma(32) stability at 10 min after shift from 30 to 42 degrees C revealed more extreme instability (tr(1/2)=20 s) th an had previously been thought. Thus, the dynamic change in sigma(32) stabi lity during the heat-shock response largely accounts for the apparent shuto ff of sigma(32) synthesis observed with a longer pulse. These results sugge st a mechanism for maintaining the intricate balance between the antagonist ic pathways: the rpoH translation as determined primarily by ambient temper ature and the turnover of sigma(32) as modulated by the chaperone (and pres umably protease)-mediated autogenous control.