Substitutions in region 2.4 of σ70 allow recognition of the σS-dependent aidB promoter

Abstract

The strict dependence of transcription from the aidB promoter (PaidB) on the EσS form of RNA polymerase is because of the presence of a C nucleotide as the first residue of the -10 promoter sequence (-12C), which does not allow an open complex formation by Eσ70. In this report, σ70 mutants carrying either the Q437H or the T440I single amino acid substitutions, which allow -12C recognition by σ70, were tested for their ability to carry out transcription from PaidB. The Gln-437 and Thr-440 residues are located in region 2.4 of σ70 and correspond to Gln-152 and Glu-155 in σS. Interestingly, the Q437H mutant of σ70, but not T440I, was able to promote an open complex formation and to initiate transcription at PaidB. In contrast to T440I, a T440E mutant was proficient in carrying out transcription from PaidB. No σ70 mutant displayed significantly increased interaction with a PaidB mutant in which the -12C was substituted by a T (PaidB(C12T)), which is also efficiently recognized by wild type σ70. The effect of the T440E mutation suggests that the corresponding Glu-155 residue in σS might be involved in -12C recognition. However, substitution to alanine of the Glu-155 residue, as well as of Gln-152, in the σS protein did not significantly affect EσS interaction with PaidB. Our results reiterate the importance of the -12C residue for σS-specific promoter recognition and strongly suggest that interaction with the -10 sequence and open complex formation are carried out by different determinants in the two σ factors.