Abstract

Translation termination is the final stage of protein synthesis. It includes at least two essential functions, stop codon recognition and polypeptide chain release. In eukaryotic organisms, the class I release factor eRF1 recognizes each of the three termination codons (UAA, UAG, and UGA) and mediates release of the nascent polypeptide chain. The class II release factor eRF3 assists the termination process in a GTP-dependent manner. The long-term goal of this project is to better understand the process of translation termination so therapeutic strategies aimed at the suppression of disease-causing premature stop mutations can be developed. The eRF1 protein contains three discrete domains. A consideration of structural and genetic data led to the proposal that domain 1 mediates stop codon recognition;domain 2 interacts with the peptidyl transferase center of the ribosome to facilitate polypeptide chain release;and domain 3 mediates the interaction between eRF1 with eRF3. Competing models argue that either the TASNIKS motif or the YCF motif in domain 1 is critical for stop codon recognition.
The first aim of this proposal will identify key residues of eRF1 domain 1 involved in stop codon recognition to test the relative merits of these competing models. The yeast SUP45 gene encodes eRF1. We recently discovered that the half-life of the SUP45 mRNA is regulated by the efficiency of the termination process. This mechanism leads to an increase in the eRF1 protein level when termination is compromised.
The second aim of this proposal will test this model and explore how this novel regulatory mechanism controls SUP45 mRNA and eRF1 protein levels. We recently found that the previously uncharacterized protein Tpa1p influences the efficiency of translation termination, mRNA poly(A) tail length, and mRNA half-life in yeast cells. This led us to propose a model in which Tpa1p couples translation termination to the deadenylation of cellular mRNAs.
The third aim of this proposal will test various aspects of this model so we can better understand the important interplay between translation termination and mRNA stability.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM068854-08
Application #
7995246
Study Section
Molecular Genetics C Study Section (MGC)
Program Officer
Bender, Michael T
Project Start
2003-06-01
Project End
2012-11-30
Budget Start
2010-12-01
Budget End
2012-11-30
Support Year
8
Fiscal Year
2011
Total Cost
$298,440
Indirect Cost

  Institution

Name
University of Alabama Birmingham
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294

  Publications

Kelly, Shane P; Bedwell, David M (2015) Both the autophagy and proteasomal pathways facilitate the Ubp3p-dependent depletion of a subset of translation and RNA turnover factors during nitrogen starvation in Saccharomyces cerevisiae. RNA 21:898-910
Conard, Sara E; Buckley, Jessica; Dang, Mai et al. (2012) Identification of eRF1 residues that play critical and complementary roles in stop codon recognition. RNA 18:1210-21
Keeling, Kim M; Bedwell, David M (2011) Suppression of nonsense mutations as a therapeutic approach to treat genetic diseases. Wiley Interdiscip Rev RNA 2:837-52
Vallabhaneni, Haritha; Fan-Minogue, Hua; Bedwell, David M et al. (2009) Connection between stop codon reassignment and frequent use of shifty stop frameshifting. RNA 15:889-97
Fan-Minogue, Hua; Du, Ming; Pisarev, Andrey V et al. (2008) Distinct eRF3 requirements suggest alternate eRF1 conformations mediate peptide release during eukaryotic translation termination. Mol Cell 30:599-609
Fan-Minogue, Hua; Bedwell, David M (2008) Eukaryotic ribosomal RNA determinants of aminoglycoside resistance and their role in translational fidelity. RNA 14:148-57
Keeling, Kim M; Salas-Marco, Joe; Osherovich, Lev Z et al. (2006) Tpa1p is part of an mRNP complex that influences translation termination, mRNA deadenylation, and mRNA turnover in Saccharomyces cerevisiae. Mol Cell Biol 26:5237-48
Salas-Marco, Joe; Fan-Minogue, Hua; Kallmeyer, Adam K et al. (2006) Distinct paths to stop codon reassignment by the variant-code organisms Tetrahymena and Euplotes. Mol Cell Biol 26:438-47
Kallmeyer, Adam K; Keeling, Kim M; Bedwell, David M (2006) Eukaryotic release factor 1 phosphorylation by CK2 protein kinase is dynamic but has little effect on the efficiency of translation termination in Saccharomyces cerevisiae. Eukaryot Cell 5:1378-87
Salas-Marco, Joe; Bedwell, David M (2005) Discrimination between defects in elongation fidelity and termination efficiency provides mechanistic insights into translational readthrough. J Mol Biol 348:801-15

Showing the most recent 10 out of 12 publications