Genome-wide single-cell-level screen for protein abundance and localization changes in response to DNA damage in S. cerevisiae

Nucleic Acids Res. 2013 Nov;41(20):9310-24. doi: 10.1093/nar/gkt715. Epub 2013 Aug 9.

Abstract

An effective response to DNA damaging agents involves modulating numerous facets of cellular homeostasis in addition to DNA repair and cell-cycle checkpoint pathways. Fluorescence microscopy-based imaging offers the opportunity to simultaneously interrogate changes in both protein level and subcellular localization in response to DNA damaging agents at the single-cell level. We report here results from screening the yeast Green Fluorescent Protein (GFP)-fusion library to investigate global cellular protein reorganization on exposure to the alkylating agent methyl methanesulfonate (MMS). Broad groups of induced, repressed, nucleus- and cytoplasm-enriched proteins were identified. Gene Ontology and interactome analyses revealed the underlying cellular processes. Transcription factor (TF) analysis identified principal regulators of the response, and targets of all major stress-responsive TFs were enriched amongst the induced proteins. An unexpected partitioning of biological function according to the number of TFs targeting individual genes was revealed. Finally, differential modulation of ribosomal proteins depending on methyl methanesulfonate dose was shown to correlate with cell growth and with the translocation of the Sfp1 TF. We conclude that cellular responses can navigate different routes according to the extent of damage, relying on both expression and localization changes of specific proteins.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Chromatin Assembly and Disassembly
  • Cytoplasm / chemistry
  • DNA Damage*
  • DNA Repair*
  • Gene Expression Regulation, Fungal
  • Gene Regulatory Networks
  • Genome, Fungal
  • Lipids / biosynthesis
  • Methyl Methanesulfonate / toxicity
  • Nuclear Proteins / analysis
  • Protein Biosynthesis
  • Proteolysis
  • Ribosomal Proteins / biosynthesis
  • Ribosomal Proteins / genetics
  • Ribosomes / metabolism
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae Proteins / analysis
  • Saccharomyces cerevisiae Proteins / biosynthesis*
  • Saccharomyces cerevisiae Proteins / genetics
  • Transcription Factors / metabolism

Substances

  • Lipids
  • Nuclear Proteins
  • Ribosomal Proteins
  • Saccharomyces cerevisiae Proteins
  • Transcription Factors
  • Methyl Methanesulfonate