A proteomic analysis of ataxia telangiectasia-mutated (ATM)/ATM-Rad3-related (ATR) substrates identifies the ubiquitin-proteasome system as a regulator for DNA damage checkpoints.
ATM (ataxia telangiectasia-mutated) and ATR (ATM-Rad3-related) are proximal checkpoint kinases that regulate DNA damage response (DDR). Identification and characterization of ATM/ATR substrates hold the keys for the understanding of DDR. Few techniques are available to identify protein kinase substrates. Here, we screened for potential ATM/ATR substrates using phospho-specific antibodies against known ATM/ATR substrates. We identified proteins cross-reacting to phospho-specific antibodies in response to DNA damage by mass spectrometry. We validated a subset of the candidate substrates to be phosphorylated in an ATM/ATR-dependent manner in vivo. Combining with a functional checkpoint screen, we identified proteins that belong to the ubiquitin-proteasome system (UPS) to be required in mammalian DNA damage checkpoint control, particularly the G(1) cell cycle checkpoint, thus revealing protein ubiquitylation as an important regulatory mechanism downstream of ATM/ATR activation for checkpoint control.
Pubmed ID: 17478428 RIS Download
Amino Acid Sequence | Antibodies, Phospho-Specific | Ataxia Telangiectasia Mutated Proteins | Cell Cycle | Cell Cycle Proteins | DNA Damage | DNA Repair | DNA-Binding Proteins | HeLa Cells | Humans | Proteasome Endopeptidase Complex | Protein-Serine-Threonine Kinases | Proteome | Reproducibility of Results | Tumor Suppressor Proteins | Ubiquitin