Quantitative Proteomics Reveal ATM Kinase-dependent Exchange in DNA Damage Response Complexes
| Autor: | Christopher J. Bakkenist, Rohith Srivas, Banu Dost, Trey Ideker, Nuno Bandeira, Bennett Van Houten, Serah Choi, Thomas P. Conrads, Gregory A. Gibson, Brian L. Hood, Katherine Y. Fu, Simon C. Watkins |
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| Jazyk: | angličtina |
| Rok vydání: | 2012 |
| Předmět: |
Proteomics
DNA repair DNA damage Cell Survival Morpholines Cell Cycle Proteins Ataxia Telangiectasia Mutated Proteins Biology Protein Serine-Threonine Kinases Biochemistry Article Cell Line Stable isotope labeling by amino acids in cell culture Humans CHEK1 Protein Interaction Maps DNA-PKcs Annexin A1 Cell Proliferation Tumor Suppressor Proteins Intracellular Signaling Peptides and Proteins Fluorescence recovery after photobleaching General Chemistry Chromatin Cell biology DNA-Binding Proteins DNA Repair Enzymes Pyrones Gene Knockdown Techniques Multiprotein Complexes RNA Interference Tumor Suppressor p53-Binding Protein 1 Ataxia telangiectasia and Rad3 related DNA Damage Protein Binding |
| Popis: | ATM is a protein kinase that initiates a well-characterized signaling cascade in cells exposed to ionizing radiation (IR). However, the role for ATM in coordinating critical protein interactions and subsequent exchanges within DNA damage response (DDR) complexes is unknown. We combined SILAC-based tandem mass spectrometry and a subcellular fractionation protocol to interrogate the proteome of irradiated cells treated with or without the ATM kinase inhibitor KU55933. We developed an integrative network analysis to identify and prioritize proteins that were responsive to KU55933, specifically in chromatin, and that were also enriched for physical interactions with known DNA repair proteins. This analysis identified 53BP1 and annexin A1 (ANXA1) as strong candidates. Using fluorescence recovery after photobleaching, we found that the exchange of GFP-53BP1 in DDR complexes decreased with KU55933. Further, we found that ANXA1 knockdown sensitized cells to IR via a mechanism that was not potentiated by KU55933. Our study reveals a role for ATM kinase activity in the dynamic exchange of proteins in DDR complexes and identifies a role for ANXA1 in cellular radioprotection. |
| Databáze: | OpenAIRE |
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