p53 Binding Protein 53BP1 Is Required for DNA Damage Responses and Tumor Suppression in Mice

Abstract

53BP1 is a p53 binding protein of unknown function that binds to the central DNA-binding domain of p53.It relocates to the sites of DNA strand breaks in response to DNA damage and is a putative substrate of the ataxia telangiectasia-mutated (ATM) kinase. To study the biological role of 53BP1, we disrupted the 53BP1 gene in the mouse. We show that, similar to ATM ؊/؊ mice, 53BP1-deficient mice were growth retarded, immune deficient, radiation sensitive, and cancer prone. 53BP1؊/؊ cells show a slight S-phase checkpoint defect and prolonged G 2 /M arrest after treatment with ionizing radiation. Moreover, 53BP1؊/؊ cells feature a defective DNA damage response with impaired Chk2 activation. These data indicate that 53BP1 acts downstream of ATM and upstream of Chk2 in the DNA damage response pathway and is involved in tumor suppression.Defects in DNA damage recognition and repair mechanisms are associated with cancer predisposition. The tumor suppressor protein p53, a sequence specific transcription factor, plays a central role in the response of mammalian cells to genotoxic stress. 53BP1 (p53 binding protein 1) was cloned as a protein that interacts with the DNA-binding domain of p53 (13). It contains a tandem BRCT (BRCA1 C terminus) motif (5) with sequence homology to the tumor suppressor BRCA1 and DNA damage checkpoint protein scRad9. 53BP1 binds through the first of its C-terminal BRCT repeats and the inter-BRCT linker region to the central DNA-binding domain of p53 (7,15) and has been shown to enhance p53-mediated transcription of reporter genes (14). More recently, in vitro studies suggest that 53BP1 participates in the cellular response to DNA damage. 53BP1 relocates to multiple nuclear foci within minutes after exposure of cells to ionizing radiation (IR) (2,22,23,28). These foci colocalize with known DNA damage response proteins such as phosphorylated H2AX, Rad50/ Mre11/NBS1, BRCA1, and Rad51 at sites of DNA lesions (2,22,23). 53BP1 becomes hyperphosphorylated in response to IR, and several lines of evidence suggest that 53BP1 is a downstream target of the ataxia telangiectasia-mutated (ATM) kinase, the product of the gene mutated in ataxia telangiectasia (2,22,28). Furthermore, 53BP1 localizes to kinetochores in mitotic cells, suggesting a potential function of 53BP1 in mitotic checkpoint signaling (16).To study the biological function of 53BP1 in mammals, we created 53BP1-deficient mice. We report here that mice lacking 53BP1 are viable and display a phenotype that partially overlaps with that of ATM-deficient mice. 53BP1-deficient mice are growth retarded, immune deficient, radiation sensitive, and cancer prone. Thus, 53BP1 is required for an appropriate cellular response to DNA damage in vivo. MATERIALS AND METHODSGene targeting and generation of 53BP1-deficient mice. A mouse 53BP1 cDNA fragment was used as a probe to isolate 53BP1 mouse genomic DNA from a mouse 129 genomic DNA phage library (Stratagene). The genomic DNA was cloned into pZErO-2 (Invitrogen), and the exon-intron structure characterized by ...

Integrative analysis reveals 53BP1 copy loss and decreased expression in a subset of human diffuse large B-cell lymphomas

“…Although previous murine genetic models strongly implicated 53bp1 copy loss in lymphomagenesis, the current study is the first report of 53BP1 hemizygosity in primary human lymphomas. In conjunction with murine analyses of 53bp1 þ /À cells, which have reduced levels of the damage response protein and increased chromosomal breakage and aneuploidy (Ward et al, 2003b(Ward et al, , 2005, the current human studies suggest that 53BP1 haploinsufficiency may be a predisposing event in certain DLBCLs. Like other 'caretaker' genes involved in DNA repair, 53BP1 likely functions in a dose-dependent manner with haploinsufficiency leading to increased genomic instability, additional somatic mutations and lymphoid transformation (Fodde and Smits, 2002).…”

“…In initial murine models, 53bp1 À/À animals had an increased incidence of early-onset thymic lymphomas (Ward et al, 2003b). In these studies, there was a close association between 53bp1 protein abundance and gene copy number, with 53bp1 þ /À cells from hemizygous animals expressing intermediate levels of the damage response protein (Ward et al, 2003b). With additional long-term follow-up, both 53bp1 À/À and 53bp1 þ /À mice had an increased incidence of late-onset tumors (Ward et al, 2005).…”

DNA repair and the immune system: From V(D)J recombination to aging lymphocytes

“…Interestingly, the adverse effect of ROS on HSC and T cell development is reverted by antioxidant agents [82,86]. Mice deficient for other DNA repair factors, which are not directly involved in V(D)J recombination, such as H2AX or 53BP1, present with variable lymphopenia ( [87][88][89][90] and our unpublished observations). Whether DNA damage generated by ROS contributes to the immune defect observed in these models is certainly an interesting issue to pursue.…”

BRCA1, PARP, and 53BP1: conditional synthetic lethality and synthetic viability

“…53BP1 is a nuclear protein that contains Tudor domains and paired BRCT repeats, and is found in all mammalian cells. Upon the induction of DNA DSBs, 53BP1 rapidly redistributes from a diffuse nuclear localization to discrete foci that co-localize with phosphorylated histone H2AX and other repair proteins including BRCA1 (Schultz et al, 2000;Wang et al, 2002;Ward et al, 2003). This localization is dependent upon interaction of the Tudor domains with methylated histone residues (Huyen et al, 2004;Sanders et al, 2004).…”