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Published 2008 | public
Book Section - Chapter

Chapter 1: Methods to Study No-Go mRNA Decay in Saccharomyces cerevisiae

Abstract

In eukaryotic cells, conserved mRNA surveillance systems target and degrade aberrant mRNAs, eliminating translation errors that occur during protein synthesis and thereby imposing quality control of gene expression. Two such cytoplasmic quality control systems, nonsense-mediated mRNA decay and nonstop mRNA decay, have evolved to target mRNAs with aberrancies in translation. A third novel quality control system has been identified for yeast mRNAs with defects in translation elongation due to strong translation pause sites. This subset of mRNAs with ribosome pause sites is recognized and targeted for degradation by an endonucleolytic cleavage in a process referred to as no-go mRNA decay (NGD). The methods described herein are designed to aid in the study of NGD in Saccharomyces cerevisiae. They include procedures to create an efficient translation elongation pause, assay decay characteristics of NGD substrates, and characterize NGD-dependent endonucleolytic cleavage of mRNA. The logic of the design and methods described can be modulated and used for the identification and analysis of novel RNA quality control pathways in other organisms.

Additional Information

© 2008 Elsevier. Available online 10 February 2009. I thank Roy Parker and Kristian Baker for discussions and critical review of the manuscript. All work described in this chapter has been done at the Department of Molecular and Cellular Biology and HHMI, University of Arizona, Tucson, Arizona, and supported by funds from the Howard Hughes Medical Institute and the National Institute of Health.

Additional details

Created:
August 22, 2023
Modified:
January 12, 2024