The global transcriptional response of Escherichia coli to induced σ32 protein involves σ32 regulon activation followed by inactivation and degradation of σ32 in vivo

sigma(32) is the first alternative sigma factor discovered in Escherichia coli and can direct transcription of many genes in response to heat shock stress. To define the physiological role of sigma(32), we have used transcription profiling experiments to identify, on a genome-wide basis, genes under the control of sigma(32) in E. coli by moderate induction of a plasmid-borne rpoH gene under defined, steady-state growth conditions. Together with a bioinformatics approach, we successfully confirmed genes known previously to be directly under the control of sigma(32) and also assigned many additional genes to the sigma(32) regulon. In addition, to understand better the functional relevance of the increased amount of sigma(32) to changes in the transcriptional level of sigma(32)-dependent genes, we measured the protein level of sigma(32) both before and after induction by a newly developed quantitative Western blot method. At a normal constant growth temperature (37 degrees C), we found that the sigma(32) protein level rapidly increased, plateaued, and then gradually decreased after induction, indicating sigma(32) can be regulated by genes in its regulon and that the mechanisms of sigma(32) synthesis, inactivation, and degradation are not strictly temperature-dependent. The decrease in the transcriptional level of sigma(32)-dependent genes occurs earlier than the decrease in full-length sigma(32) in the wild type strain, and the decrease in the transcriptional level of sigma(32)-dependent genes is greatly diminished in a Delta DnaK strain, suggesting that DnaK can act as an anti-sigma factor to functionally inactivate sigma(3)2 and thus reduce sigma(32)-dependent transcription in vivo.