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Using the E1A Minigene Tool to Study mRNA Splicing Changes
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Engineering ER-stress dependent non-conventional mRNA splicing.

Weihan Li1,2, Voytek Okreglak1,2, Jirka Peschek1,2

  • 1Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States.

Elife
|July 10, 2018
PubMed
Summary
This summary is machine-generated.

The endoplasmic reticulum kinase/RNase Ire1 has specialized functions in yeast, with distinct RNase specificities and RNA structures separating mRNA splicing from decay. Researchers engineered both functions into one yeast strain.

Keywords:
RNA processingS. cerevisiaeS. pombebiochemistrycell biologychemical biologyevolutionary biologynon-conventional mRNA splicingunfolded protein response

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Area of Science:

  • Molecular Biology
  • Cellular Biology
  • Yeast Genetics

Background:

  • The endoplasmic reticulum (ER) maintains protein homeostasis by balancing protein folding capacity and burden.
  • The ER kinase/RNase Ire1 is crucial for ER protein homeostasis, regulating gene expression via mRNA splicing and controlling protein burden via mRNA decay.
  • In yeast, these two Ire1 functions have diverged between species.

Purpose of the Study:

  • To investigate the evolutionary specialization of Ire1 orthologs in yeast.
  • To understand the molecular basis for the separation of Ire1's splicing and decay functions.
  • To engineer dual Ire1 functionality into a single yeast species.

Main Methods:

  • Comparative analysis of Ire1 orthologs in *Saccharomyces cerevisiae* and *Schizosaccharomyces pombe*.
  • Investigation of RNase specificities and RNA structural features of Ire1 substrates.
  • Genetic engineering of *S. pombe* Ire1 to create a dual-function enzyme.

Main Results:

  • Ire1 orthologs in different yeast species have specialized RNase specificities, leading to distinct functional outputs.
  • Specific RNA structural features differentiate mRNA substrates targeted for splicing versus decay by Ire1.
  • Engineered *S. pombe* Ire1 gained the ability to perform both mRNA splicing and decay functions.

Conclusions:

  • Evolutionary divergence of RNase specificity and substrate recognition drives the specialization of Ire1 functions in yeast.
  • Understanding these mechanisms allows for the engineering of novel Ire1 activities.
  • This work provides insights into the regulation of ER protein homeostasis and potential therapeutic targets.