The reduction in sigma-promoter recognition flexibility as induced by core RNAP is required for sigma to discern the optimal promoter spacing

Abstract

Sigma (σ) factors are bacterial transcription initiation factors that direct transcription at cognate promoters. The promoters recognized by primary σ are composed of -10 and -35 consensus elements separated by a spacer of 17±1 bp for optimal activity. However, how the optimal promoter spacing is sensed by the primary σ remains unclear. In the present study, we examined this issue using a transcriptionally active Bacillus subtilis N-terminally truncated σA (SND100-σA). The results of the present study demonstrate that SND100-σA binds specifically to both the -10 and -35 elements of the trnS spacing variants, of which the spacer lengths range from 14 to 21 bp, indicating that simultaneous and specific recognition of promoter -10 and -35 elements is insufficient for primary σ to discern the optimal promoter spacing. Moreover, shortening in length of the flexible linker between the two promoter DNA-binding domains of σA also does not enable SND100-σA to sense the optimal promoter spacing. Efficient recognition of optimal promoter spacing by SND100-σA requires core RNAP (RNA polymerase) which reduces the flexibility of simultaneous and specific binding of SND100-σA to both promoter -10 and -35 elements. Thus the discrimination of optimal promoter spacing by σ is core-dependent.

The core‐independent promoter‐specific binding of Bacillus subtilis σB

“…The inability of the primary σ factor to specifically recognize promoter DNA has been ascribed to an indirect steric and/or electrostatic mechanism , or the close proximity of the negatively charged σ 1.1 to the positively charged promoter recognition domains of the σ factor, which makes the σ factor adopt a compact structure that is incompatible with DNA binding . Consistent with this possibility, a Bacillus subtilis N‐terminally truncated primary σ A (SND100‐σ A ) lacking σ 1.1 is able to core‐independently and specifically recognize both the −10 and −35 elements of promoters despite an inability to discern the optimal promoter spacing . Core‐independent promoter −10 element‐specific binding activity has been reported for the alternative group 3 σ factor, σ D of B. subtilis and Aeribacillus pallidus AC6, which also lack σ 1.1 .…”

Determinants of mer Promoter Activity from Pseudomonas aeruginosa

“…To provide the possibility that the base at position −13 interacts with the RNA polymerase holoenzyme, we performed a crystal structure analysis. The most conserved region 2 (σ 2 ) of sigma factor 70 (σ 70 ) was divided into four subregions; subregion 2.3 (σ 2.3 ) is implicated in DNA melting, and subregion 2.4 (σ 2.4 ) is associated with recognizing the −10 element [54,75]. An α-helix containing a sequence 430 YATWWIRQAITRSIAD 445 in σ 2.3 to σ 2.4 interacts with the −10 element of the promoter.…”

“…In addition, some studies support an optimal spacer length of 17 bp [48][49][50], while others suggest an optimal spacer length of 16-18 bp [51-53]. Typically, even if the spacer differs from the optimum by only 1 bp, the promoter activity is significantly reduced [54][55][56][57][58]. However, a P mer derivative with a 17 bp spacer exhibited constitutive activity similar to that of derivatives with 18 bp spacers in previous in vivo studies [27].…”