Anti-pausing activity of region 4 of the RNA polymerase σ subunit and its regulation by σ-remodeling factors

Abstract

The basal transcription factors of cellular RNA polymerases (RNAPs) stimulate the initial RNA synthesis via poorly understood mechanisms. Here, we explored the mechanism employed by the bacterial factor {sigma} in promoter-independent initial transcription. We found that the RNAP holoenzyme lacking the promoter-binding domain {sigma}4 is ineffective in de novo transcription initiation and displays high propensity to pausing upon extension of RNAs 3 to 7 nucleotides in length. The {sigma}4 domain stabilizes short RNA:DNA hybrids and suppresses pausing by stimulating RNAP active-center translocation. The anti-pausing activity of {sigma}4 is modulated by its interaction with the {beta} subunit flap domain and by the {sigma} remodeling factors AsiA and RbpA. Our results suggest that the presence of {sigma}4 within the RNA exit channel compensates for the intrinsic instability of short RNA:DNA hybrids by increasing RNAP processivity, thus favoring productive transcription initiation. This "RNAP boosting" activity of the initiation factor is shaped by the the thermodynamics of RNA:DNA interactions and thus, should be relevant for any factor-dependent RNAP.