Potential role for 53BP1 in DNA end-joining repair through direct interaction with DNA.
Upon DNA damage, p53-binding protein 1 (53BP1) relocalizes to sites of DNA double-strand breaks and forms discrete nuclear foci, suggesting its role in DNA damage responses. We show that 53BP1 changed its localization from the detergent soluble to insoluble fraction after treatment of cells with x-ray, but not with ultraviolet or hydroxyurea. Either DNase or phosphatase treatment of the insoluble fraction released 53BP1 into the soluble fraction, showing that 53BP1 binds to chromatin in a phosphorylation-dependent manner after X-irradiation of cells. 53BP1 was retained at discrete nuclear foci in X-irradiated cells even after detergent extraction of cells, showing that the chromatin binding of 53BP1 occurs at sites of DNA double-strand breaks. The minimal domain for focus formation was identified by immunofluorescence staining of cells ectopically expressed with 53BP1 deletion mutants. This domain consisted of conserved Tudor and Myb motifs. The Tudor plus Myb domain possessed chromatin binding activity in vivo and bound directly to both double-stranded and single-stranded DNA in vitro. This domain also stimulated end-joining by DNA ligase IV/Xrcc4, but not by T4 DNA ligase in vitro. We conclude that 53BP1 has the potential to participate directly in the repair of DNA double-strand breaks.
Iwabuchi
Kuniyoshi
K
Department of Biochemistry, Kanazawa Medical University, 1-1 Daigaku, Uchinada, Kahoku-gun, Ishikawa 920-0293, Japan.
Basu
Balaka Piku
BP
Kysela
Boris
B
Kurihara
Takayuki
T
Shibata
Masao
M
Guan
Deyu
D
Cao
Yongheng
Y
Hamada
Tomio
T
Imamura
Kouji
K
Jeggo
Penny A
PA
Date
Takayasu
T
Doherty
Aidan J
AJ
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004249:DNA Damage; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000737:chemistry; D004260:DNA Repair; D003902:Detergents / Q000494:pharmacology; D004307:Dose-Response Relationship, Radiation; D017353:Gene Deletion; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D047908:Intracellular Signaling Peptides and Proteins; D018386:Kinetochores / Q000737:chemistry; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D020626:Oncogene Proteins v-myb / Q000737:chemistry; D010750:Phosphoproteins; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D000071857:Tumor Suppressor p53-Binding Protein 1
D002352:Carrier Proteins; D002843:Chromatin; D003902:Detergents; D047908:Intracellular Signaling Peptides and Proteins; C000607893:LIG4 protein, human; D020626:Oncogene Proteins v-myb; D010750:Phosphoproteins; C498301:TP53BP1 protein, human; D000071857:Tumor Suppressor p53-Binding Protein 1; D004247:DNA; D005982:Glutathione Transferase; D010744:Phosphoric Monoester Hydrolases; D011088:DNA Ligases; D000072481:DNA Ligase ATP