UMMS Affiliation

Department of Microbiology and Physiological Systems

Publication Date

5-31-2018

Document Type

Article

Disciplines

Biochemistry, Biophysics, and Structural Biology | Cell Biology | Cellular and Molecular Physiology

Abstract

Termination of RNA Polymerase II (Pol II) activity serves a vital cellular role by separating ubiquitous transcription units and influencing RNA fate and function. In the yeast Saccharomyces cerevisiae, Pol II termination is carried out by cleavage and polyadenylation factor (CPF-CF) and Nrd1-Nab3-Sen1 (NNS) complexes, which operate primarily at mRNA and non-coding RNA genes, respectively. Premature Pol II termination (attenuation) contributes to gene regulation, but there is limited knowledge of its prevalence and biological significance. In particular, it is unclear how much crosstalk occurs between CPF-CF and NNS complexes and how Pol II attenuation is modulated during stress adaptation. In this study, we have identified an attenuator in the DEF1 DNA repair gene, which includes a portion of the 5'-untranslated region (UTR) and upstream open reading frame (ORF). Using a plasmid-based reporter gene system, we conducted a genetic screen of 14 termination mutants and their ability to confer Pol II read-through defects. The DEF1 attenuator behaved as a hybrid terminator, relying heavily on CPF-CF and Sen1 but without Nrd1 and Nab3 involvement. Our genetic selection identified 22 cis-acting point mutations that clustered into four regions, including a polyadenylation site efficiency element that genetically interacts with its cognate binding-protein Hrp1. Outside of the reporter gene context, a DEF1 attenuator mutant increased mRNA and protein expression, exacerbating the toxicity of a constitutively active Def1 protein. Overall, our data support a biologically significant role for transcription attenuation in regulating DEF1 expression, which can be modulated during the DNA damage response.

Keywords

Def1, Nrd1/Nab3/Sen1 complex (NNS), RNA polymerase II termination, attenuation, cleavage and polyadenylation complex (CPF-CF)

Rights and Permissions

Copyright © 2018 Whalen et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

DOI of Published Version

10.1534/g3.118.200072

Source

G3 (Bethesda). 2018 May 31;8(6):2043-2058. doi: 10.1534/g3.118.200072. Link to article on publisher's site

Journal/Book/Conference Title

G3 (Bethesda, Md.)

Related Resources

Link to Article in PubMed

PubMed ID

29686108

Creative Commons License

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.

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