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

Abstract

σ32 is the first alternative σ factor discovered in Escherichia coli and can direct transcription of many genes in response to heat shock stress. To define the physiological role of σ32, we have used transcription profiling experiments to identify, on a genome-wide basis, genes under the control of σ32 in E. coli by moderate induction of a plasmid-bome 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 σ32 and also assigned many additional genes to the σ32 regulon. In addition, to understand better the functional relevance of the increased amount of σ32 to changes in the transcriptional level of σ32-dependent genes, we measured the protein level of σ32 both before and after induction by a newly developed quantitative Western blot method. At a normal constant growth temperature (37 °C), we found that the σ32 protein level rapidly increased, plateaued, and then gradually decreased after induction, indicating σ32 can be regulated by genes in its regulon and that the mechanisms of σ32 synthesis, inactivation, and degradation are not strictly temperature-dependent. The decrease in the transcriptional level of σ32-dependent genes occurs earlier than the decrease in full-length σ32 in the wild type strain, and the decrease in the transcriptional level of σ32-dependent genes is greatly diminished in a ΔDnaK strain, suggesting that DnaK can act as an anti-σ factor to functionally inactivate σ32 and thus reduce σ32-dependent transcription in vivo. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.