Yeast DEAD Box Protein Mss116p Is a Transcription Elongation Factor That Modulates the Activity of Mitochondrial RNA Polymerase

Abstract

DEAD box proteins have been widely implicated in regulation of gene expression. Here, we show that the yeast Saccharomyces cerevisiae DEAD box protein Mss116p, previously known as a mitochondrial splicing factor, also acts as a transcription factor that modulates the activity of the single-subunit mitochondrial RNA polymerase encoded by RPO41. Binding of Mss116p stabilizes paused mitochondrial RNA polymerase elongation complexes in vitro and favors the posttranslocated state of the enzyme, resulting in a lower concentration of nucleotide substrate required to escape the pause; this mechanism of action is similar to that of elongation factors that enhance the processivity of multisubunit RNA polymerases. In a yeast strain in which the RNA splicing-related functions of Mss116p are dispensable, overexpression of RPO41 or MSS116 increases cell survival from colonies that were exposed to low temperature, suggesting a role for Mss116p in enhancing the efficiency of mitochondrial transcription under stress conditions.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.00160-14.

ACKNOWLEDGMENTS

We are especially grateful to Mark Del Campo and Alan M. Lambowitz (University of Texas at Austin, TX) for providing purified CYT-19, Ded1p, wild-type Mss116p protein, and its mutant derivatives, the S. cerevisiae Δmss116 I⁰ strain, a plasmid construct for expression of maltose binding protein-Mss116 fusion in E. coli, and helpful discussions and comments on an early version of the manuscript. We also thank Smita Patel (RWJMS/Rutgers University, Piscataway, NJ) for helpful discussions and comments on an early version of the manuscript. We thank Sergei Borukhov for E. coli RNA polymerase preparations, assistance with in vitro transcription assays, and insightful comments; Dmitry Temiakov and Michael Anikin for helpful comments; and Ray Castagna for technical assistance.

This work was supported by grants from the National Institutes of Health (GM31847) and the New Jersey Health Foundation to W.T.M.