The stress sigma factor of RNA polymerase RpoS/σS is a solvent-exposed open molecule in solution

Abstract

In bacteria, one primary and multiple alternative sigma (σ) factors associate with the RNA polymerase core enzyme (E) to form holoenzymes (Eσ) with different promoter recognition specificities. The alternative σ factor RpoS/σ is produced in stationary phase and under stress conditions and reprograms global gene expression to promote bacterial survival. To date, the three-dimensional structure of a full-length free σ factor remains elusive. The current model suggests that extensive interdomain contacts in a free σ factor result in a compact conformation that masks the DNA-binding determinants of σ, explaining why a free σ factor does not bind double-stranded promoter DNA efficiently. Here, we explored the solution conformation of σ using amide hydrogen/deuterium exchange coupled with mass spectrometry, NMR, analytical ultracentrifugation and molecular dynamics. Our data strongly argue against a compact conformation of free σ Instead, we show that σ adopts an open conformation in solution in which the folded σ and σ domains are interspersed by domains with a high degree of disorder. These findings suggest that E binding induces major changes in both the folding and domain arrangement of σ and provide insights into the possible mechanisms of regulation of σ activity by its chaperone Crl.

Crl activates transcription by stabilizing active conformation of the master stress transcription initiation factor

“…Consistent with their remote locations in the cryo-EM structure, helix α1 from σ 1.2 (residues 56–67) and the σ 3.2 linker (residues 218–245) generally appear to be solvent-exposed in the HDX-MS experiment (Figure 3C; Figure 3—figure supplements 1 and 3). The HDX profile of σ S in the absence of Crl also agrees well with a recent report (Cavaliere et al, 2018).…”

Crl activates transcription by stabilizing the active conformation of the master stress factor σS

“…Consistent with their remote locations in the cryo-EM structure, helix α1 from σ 1.2 (residues 56-67) and σ 3.2 linker (residues 218-245) generally appear to be solvent-exposed in HDX-MS experiment ( Fig.3C and Fig.S5, S7). The HDX profile of σ S in the absence of Crl also agrees well with a recent report (Cavaliere et al, 2018).…”

Diversity, versatility and complexity of bacterial gene regulation mechanisms: opportunities and drawbacks for applications in synthetic biology

“…In contrast to the housekeeping σ 0 factor, σ S was recently shown to adopt an open conformation in solution in which the folded σ 2 and σ 4 domains are interspersed by domains with a high degree of disorder (Cavaliere et al. 2018). This open configuration of σ S also provides insight into a possible mechanism for regulation of its activity by the chaperone Crl (see below).…”