Heat induction of θ³²synthesis mediated by mRNA secondary structure: a primary step of the heat shock response in<i>Escherichia coli</i>

Induction of heat shock proteins following transfer of E. coli cells from 30 degrees C to 42 degrees C depends on rapid accumulation of sigma 32, a minor sigma factor specifically required for transcription of heat shock genes. The synthesis of sigma 32 is induced by enhancing translation of its mRNA transcribed from the rpoH (htpR) gene. We previously showed that the translational control of rpoH-lacZ gene fusion is mediated by two cis-acting rpoH coding regions presumably involving mRNA secondary structure. To further examine this model, we constructed and analyzed a set of gene fusions carrying base substitution(s) or internal deletions within rpoH, including constitutive mutations predicted to destroy the mRNA secondary structure and compensatory second-site mutations that may restore the secondary structure. The results demonstrate that base pairings between the translation initiation region of some 20 nucleotides and part of the internal complementary sequences are critical for maintaining repression during steady-state growth and for modulating heat-induced synthesis of sigma 32-beta-galactosidase fusion protein upon temperature upshift. Furthermore, some of the compensatory mutations resulted in super-repressed (non-inducible) phenotypes, suggesting that the heat induction depends on a specific nucleotide sequence(s) as well as the mRNA secondary structure within the 5'-proximal regulatory segment of rpoH coding region.