Conserved mechanisms of transcriptional pausing regulate diverse RNA polymerases

Abstract

Transcription of DNA into RNA requires multi‐subunit RNA polymerases (RNAPs) in all forms of life. Pausing is an important regulatory feature of RNAPs that facilitates formation of RNA structures and the process of termination in bacteria. The elemental pause is an offline state of RNAP caused by conformational rearrangements and may be a precursor to longer‐lived backtracked and RNA hairpin‐stabilized pausing. While the backtrack pause has been well‐characterized in eukaryotes, biochemical data describing the elemental pause are lacking and hairpin‐stabilized pausing has not been described. As these last two types of pausing are important for the proper regulation of transcription in prokaryotes, it is important to know whether they are conserved in eukaryotes. Using in vitro transcription assays on synthetic oligonucleotide scaffolds, we investigated the response of Saccharomyces cerevisiae RNA polymerase II (Pol II) to the consensus elemental pause and his hairpin‐stabilized pause sequences from Escherichia coli. We demonstrate that RNA duplexes can form within the Pol II elongation complex and modulate pausing behavior. These duplexes can also halt transcription elongation in a manner analogous to intrinsic terminator hairpins found in prokaryotes. We also characterize the previously‐described HIV‐1 TAR pause as a consensus elemental pause sequence, confirming that features of the elemental pause are conserved from bacteria to mammals. These data show that aspects of pausing as a form of transcriptional regulation are conserved from bacteria to mammals, and guide the search for novel examples of elemental and hairpin‐modulated pauses in eukaryotes.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.