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Analysis of nonsense-mediated mRNA decay in Saccharomyces cerevisiae

Bessie W Kebaara1, Kristian E Baker, Krista D Patefield

  • 1Department of Biology, Baylor University, Waco, Texas, USA.

Current Protocols in Cell Biology
|March 17, 2012
PubMed

Insights

Nonsense-mediated mRNA decay (NMD) removes faulty mRNAs with premature stops. This study details methods to analyze NMD targets, including mRNA structure and decay rates in yeast.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Nonsense-mediated mRNA decay (NMD) is a crucial cellular surveillance pathway.
  • NMD eliminates aberrant messenger RNAs (mRNAs) containing premature termination codons (PTCs).
  • This process prevents the production of truncated and potentially harmful proteins.

Purpose of the Study:

  • To provide a comprehensive set of techniques for studying the NMD pathway.
  • To enable the analysis of mRNA decay rates and structural features of NMD targets.
  • To facilitate in vivo studies of nonsense and allosuppression in Saccharomyces cerevisiae.

Main Methods:

  • Monitoring steady-state mRNA levels of NMD targets.
  • Measuring mRNA decay rates under various conditions.
  • Analyzing mRNA structural elements: cap status, 5' and 3' untranslated region (UTR) lengths, and poly(A) tail length.
  • In vivo analysis of nonsense and allosuppression in yeast.

Main Results:

  • Established protocols for quantifying NMD activity.
  • Characterized the impact of mRNA structural features on NMD efficiency.
  • Demonstrated the utility of yeast models for NMD research.

Conclusions:

  • The described methods offer a robust framework for investigating NMD.
  • Understanding NMD is vital for comprehending gene regulation and disease mechanisms.
  • This unit serves as a valuable resource for researchers studying mRNA quality control.

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