53BP1 exchanges slowly at the sites of DNA damage and appears to require RNA for its association with chromatin

Fiona Pryde, Shirin Khalili, Kathryn Robertson, Jim Selfridge, Ann-Marie Ritchie, David W Melton, Denis Jullien, Yasuhisa Adachi

Research output: Contribution to journalArticlepeer-review


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.

Original languageEnglish
Pages (from-to)2043-55
Number of pages13
JournalJournal of Cell Science
Issue numberPt 9
Publication statusPublished - 1 May 2005


  • Amino Acid Sequence
  • Animals
  • Chromatin
  • DNA Damage
  • DNA Repair
  • Fibroblasts
  • Fluorescence Recovery After Photobleaching
  • Green Fluorescent Proteins
  • HeLa Cells
  • Homozygote
  • Humans
  • Immunoprecipitation
  • Intracellular Signaling Peptides and Proteins
  • Mice
  • Mice, Transgenic
  • Microscopy, Fluorescence
  • Molecular Sequence Data
  • Mutation
  • Phosphoproteins
  • Plasmids
  • Protein Binding
  • Protein Structure, Tertiary
  • RNA
  • Ribonuclease, Pancreatic
  • Ribonucleases
  • Signal Transduction
  • Time Factors

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