S1219 residue of 53BP1 is phosphorylated by ATM kinase upon DNA damage and required for proper execution of DNA damage response.
53BP1 is phosphorylated by the protein kinase ATM upon DNA damage. Even though several ATM phosphorylation sites in 53BP1 have been reported, those sites have little functional implications in the DNA damage response. Here, we show that ATM phosphorylates the S1219 residue of 53BP1 in vitro and that the residue is phosphorylated in cells exposed to ionizing radiation (IR). Transfection with siRNA targeting ATM abolished IR-induced phosphorylation at this residue, supporting the theory that this process is mediated by the kinase. To determine the functional relevance of this phosphorylation event, a U2OS cell line expressing S1219A mutant 53BP1 was established. IR-induced foci formation of MDC1 and gammaH2AX, DNA damage signaling molecules, was reduced in this cell line, implying that S1219 phosphorylation is required for recruitment of these molecules to DNA damage sites. Furthermore, overexpression of the mutant protein impeded IR-induced G2 arrest. In conclusion, we have shown that S1219 phosphorylation by ATM is required for proper execution of DNA damage response.
Lee
Haemi
H
Reseach Institute, National Cancer Center, 111 Jungbalsan-ro, Ilsandong-gu, Goyang, Gyeonggi, 410-769, Republic of Korea.
Kwak
Hee-Jin
HJ
Cho
Il-taeg
IT
Park
Seok Hee
SH
Lee
Chang-Hun
CH
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011839:Radiation, Ionizing; D012694:Serine / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000071857:Tumor Suppressor p53-Binding Protein 1
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D047908:Intracellular Signaling Peptides and Proteins; C413163:MDC1 protein, human; D009687:Nuclear Proteins; C498301:TP53BP1 protein, human; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D000071857:Tumor Suppressor p53-Binding Protein 1; D012694:Serine; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases