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Measurement of mRNA Decay Rates in Saccharomyces cerevisiae Using rpb1-1 Strains
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Role of cytoplasmic deadenylation and mRNA decay factors in yeast apoptosis

Kalidindi K Raju1, Sumathi Natarajan2, N Sunil Kumar3

  • 1Department of Biotechnology, GRIET, Hyderabad 500090, Telangana, India kalidindirj@gmail.com.

FEMS Yeast Research
|February 13, 2015
PubMed

Insights

Defects in mRNA decay pathways in Saccharomyces cerevisiae trigger apoptosis and aging. Disruptions in decapping, exosome function, or deadenylation lead to cell death markers and altered gene expression.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Genetics

Background:

  • The 5' to 3' mRNA decay pathway is crucial for cellular regulation.
  • Dysregulation of mRNA decay is linked to apoptosis and aging in Saccharomyces cerevisiae.
  • Specific factors like DCP2, LSM1, SKI2, CCR4, and PAN2 play key roles in mRNA processing.

Purpose of the Study:

  • To investigate the role of specific mRNA decay factors in yeast apoptosis.
  • To analyze the impact of defects in decapping, exosome function, and deadenylation on cellular aging.
  • To examine the relationship between mRNA decay pathways and apoptotic markers.

Main Methods:

  • Deletion mutagenesis in Saccharomyces cerevisiae to create strains lacking specific mRNA decay factors (DCP2, LSM1, SKI2, CCR4, PAN2).
  • Assessment of apoptotic markers including reactive oxygen species levels, phosphatidylserine externalization, and chromatin fragmentation.
  • Measurement of caspase gene (YCA1) expression and protein activity.
  • Analysis of transcript levels for mRNA decapping regulators (eIF4E, Pab1).

Main Results:

  • Yeast strains lacking mRNA decapping activation factors (DCP2, LSM1), cytoplasmic exosome function (SKI2), or cytoplasmic deadenylases (CCR4, PAN2) exhibited hallmarks of apoptosis.
  • Increased reactive oxygen species, phosphatidylserine externalization, chromatin fragmentation, and enhanced YCA1 expression/activity were observed.
  • Transcript levels of eIF4E and Pab1 were elevated in deadenylation-defective strains and reduced in strains lacking exosome or decapping functions.
  • lsm1Δ and ccr4Δpan2Δ mutants showed the most pronounced apoptotic phenotypes, followed by DCP2 and SKI2 mutants.

Conclusions:

  • Defects in key mRNA decay pathways, including decapping, exosome function, and deadenylation, induce caspase-dependent apoptosis in yeast.
  • Specific mRNA decay factor deficiencies differentially impact apoptotic phenotypes and associated gene expression.
  • The study highlights the critical role of mRNA decay in maintaining cellular homeostasis and preventing premature aging.
Keywords:
apoptosisdeadenylasesdecappingexosomeflow-cytometerfluorescent microscopymRNA degradationmetacaspasereal time-PCRyeast

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