Concerted Activities of Distinct H4K20 Methyltransferases at DNA Double-Strand Breaks Regulate 53BP1 Nucleation and NHEJ-Directed Repair

Abstract

SUMMARY Although selective binding of 53BP1 to dimethylated histone H4 lysine 20 (H4K20me2) at DNA double strand breaks (DSBs) is a necessary and pivotal determinant of non-homologous end joining (NHEJ)-directed repair, the enzymes that generate H4K20me2 at DSBs were unclear. Here we determined that the PR-Set7 monomethyltransferase (H4K20me1) regulates de novo H4K20 methylation at DSBs. Rapid recruitment of PR-Set7 to DSBs was dependent on the NHEJ Ku70 protein and necessary for NHEJ-directed repair. PR-Set7 monomethyltransferase activity was required, but insufficient, for H4K20me2 and 53BP1 nucleation at DSBs. We determined that PR-Set7-mediated H4K20me1 facilitates Suv4-20 methyltransferase recruitment and catalysis to generate H4K20me2 necessary for 53BP1 binding. The orchestrated and concerted activities of PR-Set7 and Suv4-20 were required for proficient 53BP1 nucleation and DSB repair. This report identifies PR-Set7 as an essential component of NHEJ and implicates PR-Set7 as a central determinant of NHEJ-directed repair early in mammalian DSB repair pathway choice.

Biological function and regulation of histone and non-histone lysine methylation in response to DNA damage

“…These results collectively indicate that the pre-existence of H4K20me1 by PR-Set7 is required for 53BP1 recruitment primarily in vivo. In agreement with this, PR-Set7 is recruited to localized irradiation (405 nm UV laser) sites by proliferating cell nuclear antigen (PCNA) or to DSBs induced by AsiSI/I-SceI endonuclease in U2OS cells, which is dependent on Ku70, after which H4K20 is de novo methylated and 53BP1 is recruited [131][132][133]. Interestingly, it was reported that the rapid recruitment of PR-Set7 is not sufficient for 53BP1 recruitment.…”

“…Interestingly, it was reported that the rapid recruitment of PR-Set7 is not sufficient for 53BP1 recruitment. The de novo H4K20me1 then facilitates Suv4-20 methyltransferases recruitment and the generation of H4K20me2 for 53BP1 binding [133]. However, the PR-Set7 at damage sites is unstable and degraded by PCNA-coupled CRL4Cdt2 E3 ligase complex.…”

Nuclear compartmentalization of DNA repair

“…Methylation of H4 lysine 20 (H4K20me2 and H4K20me1) can be specifically recognized by the tandem Tudor domain of 53BP1 [24,25]. Although it was originally proposed that these histone modifications are not induced upon DNA damage but rather unmasked [24], it was recently shown that H4K20me2 can be deposited de novo after DSB induction [26,27]. Similarly, RNF168-mediated ubiquitylation at H2AK15 enhances 53BP1 accumulation and favors NHEJ to HR [28].…”

DNA double strand break repair pathway choice: a chromatin based decision?

“…47 It is unclear whether or not H4K20 mono and/or dimethylation are efficiently induced at the vicinity of DSBs to recruit 53BP1. 43,[48][49][50][51][52] Interestingly, it has been suggested that DSB-induced alterations in chromatin structure could allow the specific unmasking of H4K20me1/2 at the vicinity of the break. 41,45 Similarly, DSB-induced eviction of H4K20me2 binding proteins, such as L3MBTL1 and JMJD2A/KDM4A, would also permit unmasking of the preexisting H4K20me1/ 2 marks, allowing for damage-induced 53BP1 recruitment.…”