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Chair(s): Osley, Mary Ann
(SY050) 53BP1 as a sensor of DNA double strand breaks.
Huyen, Yentram1, Zgheib, Omar1, DiTullio, Richard1, Stavridi, Elena1, Halazonetis, Thanos*,1, 1 Molecular Oncogenesis Program, Philadelphia, PA, USA
ABSTRACT- The ATM kinase is activated in response to DNA double strand breaks (DSBs) and initiates a signaling pathway, which leads to activation of the p53 tumor suppressor protein and cell cycle arrest or apoptosis. The mechanism by which ATM is activated is not well elucidated. In addition, it is not clear, if ATM is directly activated by DNA DSBs or indirectly by DNA DSB sensor proteins. We have been studying the 53BP1 protein and have shown that 53BP1 is a DNA damage checkpoint protein that localizes rapidly to sites of DNA DSBs and contributes to activation of ATM. These findings raised the possibility that 53BP1 is a DNA DSB sensor. To explore this, we studied the mechanism by which 53BP1 is recruited to sites of DNA DSBs. Deletion mutants identified a 120 amino acid domain within 53BP1 that is sufficient and necessary for localization of 53BP1 to sites of DNA DSBs. This domain is conserved in the 53BP1 orthologs in budding yeast (Rad9) and fission yeast (Rhp9/Crb2). We solved the three-dimensional structure of this domain of human 53BP1 by X-ray crystallography. The domain consists of two tandem tudor folds with a deep pocket at their interface formed by evolutionarily conserved residues. In vitro, the tandem tudor domain of 53BP1 bound histone H3 methylated on Lys79. The residues that form the walls of the conserved pocket were required for binding to methylated histone H3 and also for recruitment of 53BP1 to sites of DNA DSBs. Suppression of Dot1L, the enzyme that methylates histone H3 on Lys79 in vivo, also inhibited recruitment of 53BP1 to sites of DNA DSBs. Because methylation of histone H3 Lys79 was not enhanced in response to DNA damage, we propose that 53BP1 senses DNA DSBs indirectly through changes in higher order chromatin structure that expose the 53BP1-binding site. Taken together, our findings suggest that 53BP1 is a sensor of DNA DSBs that functions upstream of ATM.
Key words: 53BP1, ATM, histone
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