Characterisation of the sites of DNA damage-induced 53BP1 phosphorylation catalysed by ATM and ATR.
The 53BP1 tumour suppressor, an important regulator of genome stability, is phosphorylated in response to ionising radiation (IR) by the ATM protein kinase, itself an important regulator of cellular responses to DNA damage. The only known sites of phosphorylation in 53BP1 are Ser25 and/or Ser29 but 53BP1 lacking these residues is still phosphorylated after DNA damage. In this study, we use mass spectrometry-based together with bioinformatic analysis to identify novel DNA damage-regulated sites of 53BP1 phosphorylation. Several new sites were identified that conform to the consensus Ser/Thr-Gln motif phosphorylated by ATM and related kinases. Phospho-specific antibodies were raised, and were used to demonstrate ATM-dependent phosphorylation of these residues in 53BP1 after exposure of cells to IR. Surprisingly, 53BP1 was also phosphorylated on these residues after exposure of cells to UV light. In this case, 53BP1 phosphorylation did not require ATM but required ATR instead. These data reveal that 53BP1 is phosphorylated on multiple residues in response to different types of DNA damage, and that 53BP1 is regulated by ATR in response to UV-induced DNA damage.
Jowsey
Paul
P
MRC Protein Phosphorylation Unit, Sir James Black Centre, University of Dundee, Dundee, Scotland, UK.
Morrice
Nicholas A
NA
Hastie
C James
CJ
McLauchlan
Hilary
H
Toth
Rachel
R
Rouse
John
J
D000595:Amino Acid Sequence; D064007:Ataxia Telangiectasia Mutated Proteins; D002384:Catalysis; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000378:metabolism*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D000071857:Tumor Suppressor p53-Binding Protein 1; D014466:Ultraviolet Rays
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C498301:TP53BP1 protein, human; D025521:Tumor Suppressor Proteins; D000071857:Tumor Suppressor p53-Binding Protein 1; C576324:ATM protein, human; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases