Promoter-proximal elongation regulates transcription in archaea

Fabian Blombach,Katherine Smollett,Thomas Fouqueau,Dorota Matelska,Finn Werner

doi:10.1038/s41467-021-25669-2

Abstract

Recruitment of RNA polymerase and initiation factors to the promoter is the only known target for transcription activation and repression in archaea. Whether any of the subsequent steps towards productive transcription elongation are involved in regulation is not known. We characterised how the basal transcription machinery is distributed along genes in the archaeon Saccharolobus solfataricus. We discovered a distinct early elongation phase where RNA polymerases sequentially recruit the elongation factors Spt4/5 and Elf1 to form the transcription elongation complex (TEC) before the TEC escapes into productive transcription. TEC escape is rate-limiting for transcription output during exponential growth. Oxidative stress causes changes in TEC escape that correlate with changes in the transcriptome. Our results thus establish that TEC escape contributes to the basal promoter strength and facilitates transcription regulation. Impaired TEC escape coincides with the accumulation of initiation factors at the promoter and recruitment of termination factor aCPSF1 to the early TEC. This suggests two possible mechanisms for how TEC escape limits transcription, physically blocking upstream RNA polymerases during transcription initiation and premature termination of early TECs.

Highlights

Recruitment of RNA polymerase and initiation factors to the promoter is the only known target for transcription activation and repression in archaea
We mapped the genome-wide occupancy of RNA polymerase (RNAP), initiation, elongationand termination factors to shed light on how the individual stages of transcription are subject to transcription regulation in S. solfataricus
In order to obtain the resolution that separates pre-initiation complex (PIC) from promoter-proximal, early transcription elongation complex (TEC), we adapted a ChIP-exo approach for RNAP and initiation factors TFB and TFEβ that includes 5′->3′ exonuclease-trimming of the immunoprecipitated-DNA fragments[46]

Summary

Introduction

Recruitment of RNA polymerase and initiation factors to the promoter is the only known target for transcription activation and repression in archaea. Impaired TEC escape coincides with the accumulation of initiation factors at the promoter and recruitment of termination factor aCPSF1 to the early TEC. Well-established processes of postrecruitment regulation include Sigma70-dependent pausing and transcription attenuation mediated by premature termination[12,13,14]. Another possible underlying molecular mechanism might be pausing during initial transcription[15,16], though its contribution to genome-wide gene regulation remains to be investigated[17]. The latter involves termination factor aCPSF1 (or FttA)[34], a ribonuclease that is evolutionary related to the RNAP II termination factor CPSF73 and the integrator subunit Ints[11]

Methods

Results

Conclusion