title: 53BP1 exchanges slowly at the sites of DNA damage and appears to require RNA for its association with chromatin.
abstract: 53BP1 protein is re-localized to the sites of DNA damage after ionizing radiation (IR) and is involved in DNA-damage-checkpoint signal transduction. We examined the dynamics of GFP-53BP1 in living cells. The protein starts to accumulate at the sites of DNA damage 2-3 minutes after damage induction. Fluorescence recovery after photobleaching experiments showed that GFP-53BP1 is highly mobile in non-irradiated cells. Upon binding to the IR-induced nuclear foci, the mobility of 53BP1 reduces greatly. The minimum (M) domain of 53BP1 essential for targeting to IR induced foci consists of residues 1220-1703. GFP-M protein forms foci in mouse embryonic fibroblast cells lacking functional endogenous 53BP1. The M domain contains a tandem repeat of Tudor motifs and an arginine- and glycine-rich domain (RG stretch), which are often found in proteins involved in RNA metabolism, the former being essential for targeting. RNase A treatment dissociates 53BP1 from IR-induced foci. In HeLa cells, dissociation of the M domain without the RG stretch by RNase A treatment can be restored by re-addition of nuclear RNA in the early stages of post-irradiation. 53BP1 immunoprecipitates contain some RNA molecules. Our results suggest a possible involvement of RNA in the binding of 53BP1 to chromatin damaged by IR.
authors.affiliation: The Wellcome Trust Centre for Cell Biology, The Institute of Cell and Molecular Biology, The University of Edinburgh, The King's Buildings, Edinburgh, EH9 3JR, UK.
authors.forename: Fiona
authors.initials: F
authors.lastname: Pryde
authors.forename: Shirin
authors.initials: S
authors.lastname: Khalili
authors.forename: Kathryn
authors.initials: K
authors.lastname: Robertson
authors.forename: Jim
authors.initials: J
authors.lastname: Selfridge
authors.forename: Ann-Marie
authors.initials: AM
authors.lastname: Ritchie
authors.forename: David W
authors.initials: DW
authors.lastname: Melton
authors.forename: Denis
authors.initials: D
authors.lastname: Jullien
authors.forename: Yasuhisa
authors.initials: Y
authors.lastname: Adachi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002843:Chromatin / Q000737:chemistry*; D004249:DNA Damage; D004260:DNA Repair; D005347:Fibroblasts / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006720:Homozygote; D006801:Humans; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry*; D012259:Ribonuclease, Pancreatic / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D000071857:Tumor Suppressor p53-Binding Protein 1
chemical_list: D002843:Chromatin; C092537:Ifi202b protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D010750:Phosphoproteins; C498301:TP53BP1 protein, human; D000071857:Tumor Suppressor p53-Binding Protein 1; D049452:Green Fluorescent Proteins; D012313:RNA; D012260:Ribonucleases; D012259:Ribonuclease, Pancreatic