L3MBTL2 orchestrates ubiquitin signaling by dictating the sequential recruitment of RNF8 and RNF168 after DNA damage

Cells respond to cytotoxic DNA double strand breaks by recruiting DNA repair proteins to the damaged site. This recruitment is dependent on ubiquitylation of adjacent chromatin areas by E3 ubiquitin ligases such as RNF8 and RNF168. RNF8 and RNF168 are recruited sequentially to the double strand breaks. However, it is unclear what dictates the sequential order and recruits RNF168 to the DNA lesion. Here, we reveal that Lethal(3)malignant brain tumor-like protein 2 (L3MBTL2) is the missing link between RNF8 and RNF168. We found that L3MBTL2 is recruited by MDC1 and subsequently ubiquitylated by the E3 ligase RNF8. Ubiquitylated L3MBTL2, in turn, facilitates recruitment of RNF168 to the DNA lesion and promotes DNA double strand break repair. These results identify L3MBTL2 as a key target of RNF8 following DNA damage and demonstrates how the DNA damage response pathway is orchestrated by ubiquitin signaling.