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A ribosome profiling study of mRNA cleavage by the endonuclease RelE

Jae-Yeon Hwang1, Allen R Buskirk2

  • 1Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.

Nucleic Acids Research
|December 8, 2016
PubMed

Insights

RelE, a bacterial protein, precisely cleaves mRNA within ribosomes. This novel method improves ribosome profiling, revealing gene reading frames for the first time.

Area of Science:

  • Molecular Biology
  • Bacterial Genetics
  • Translational Control

Background:

  • RelE is a bacterial toxin involved in stress response and persistence.
  • It inhibits protein synthesis by cleaving messenger RNA (mRNA) at the ribosomal A site.
  • Previous studies indicated sequence specificity and ribosome-dependent activity.

Purpose of the Study:

  • To characterize the sequence specificity of RelE.
  • To investigate the global effects of RelE on translation using ribosome profiling.
  • To explore RelE as a tool for high-resolution bacterial ribosome profiling.

Main Methods:

  • Ribosome profiling to globally analyze translation.
  • Biochemical assays to study RelE activity.
  • Characterization of mRNA cleavage sites generated by RelE.

Main Results:

  • RelE exhibits sequence specificity in mRNA cleavage.
  • Ribosome profiling revealed dynamic cycles of mRNA cleavage, rescue, and initiation.
  • RelE generates precise 3' mRNA ends, enabling reading frame determination in bacteria.
  • Purified RelE can be used to generate ribosome footprints in vitro.

Conclusions:

  • RelE's precise mRNA cleavage offers a significant improvement over traditional methods like MNase digestion for ribosome profiling.
  • This technique provides unprecedented resolution and reveals the reading frame in bacterial translation.
  • RelE has broad potential applications in studying ribosome profiling across all domains of life.

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