Genome-wide single-cell-level screen for protein abundance and localization changes in response to DNA damage in S. cerevisiae.
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.
Department of Biological Engineering, Center for Environmental Health Sciences, Laboratory for Computational Biology and Biophysics, Department of Biology and The David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Aprotim
A
Mazumder
Laia Quiros
LQ
Pesudo
Siobhan
S
McRee
Mark
M
Bathe
Leona D
LD
Samson
D042002:Chromatin Assembly and Disassembly; D003593:Cytoplasm / Q000737:chemistry; D004249:DNA Damage; D004260:DNA Repair; D015966:Gene Expression Regulation, Fungal; D053263:Gene Regulatory Networks; D016681:Genome, Fungal; D008055:Lipids / Q000096:biosynthesis; D008741:Methyl Methanesulfonate / Q000633:toxicity; D009687:Nuclear Proteins / Q000032:analysis; D014176:Protein Biosynthesis; D059748:Proteolysis; D012269:Ribosomal Proteins / Q000096:biosynthesis / Q000235:genetics; D012270:Ribosomes / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism
D008055:Lipids; D009687:Nuclear Proteins; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D008741:Methyl Methanesulfonate