MODULATION OF THE ESCHERICHIA-COLI SIGMA(E) (RPOE) HEAT-SHOCK TRANSCRIPTION-FACTOR ACTIVITY BY THE RSEA, RSEB AND RSEC PROTEINS

The sigma(E) (RpoE) transcription factor of Escherichia coli regulates the expression of genes whose products are devoted to extracytoplasmi c activities. The sigma(E) regulon is induced upon misfolding of prote ins in the periplasm or the outer membrane. Similar to other alternati ve sigma factors, the activity of sigma(E) is tightly regulated in E. coli. We have previously shown that sigma(E) is positively autoregulat ed at the transcriptional level. DNA sequencing, coupled with transcri ptional analyses, have shown that sigma(E) is encoded by the first gen e of a four-gene operon. The second gene of this operon, rseA, encodes an anti-sigma(E) activity. This was demonstrated at both the genetic and biochemical levels. For example, mutations in rseA constitutively increase sigma(E) activity. Consistent with this, overproduction of Rs eA leads to an inhibitory effect on sigma(E) activity. Topological ana lysis of RseA suggests the existence of one transmembrane domain, with the N-terminal part localized in the cytoplasm. Overproduction of thi s N-terminal domain atone was shown to inhibit sigma(E) activity. Thes e observations were confirmed in vitro, because either purified RseA o r only its purified N-terminal domain inhibited transcription from E s igma(E)-dependent promoters. Furthermore, RseA and sigma(E) co-purify, and can be co-immunoprecipitated, and chemically cross-linked. The si gma(E) activity is further modulated by the products of the remaining genes in this operon, rseB and rseC. RseB is a periplasmic protein, wh ich negatively regulates sigma(E) activity and specifically interacts with the C-terminal periplasmic domain of RseA. In contrast, RseC is a n inner membrane protein that positively modulates sigma(E) activity. Most of these protein-protein interactions were verified in vivo using the yeast two-hybrid system.