Abstract
In response to DNA double-strand breaks (DSBs), repair proteins are recruited to the damaged sites. Ubiquitin signaling plays a critical role in coordinating protein recruitment during the DNA damage response. Here, we find that the microRNA biogenesis factor DGCR8 promotes tumor resistance to X-ray radiation independently of its Drosha-binding ability. Upon radiation, the kinase ATM and the deubiquitinase USP51 mediate the activation and stabilization of DGCR8 through phosphorylation and deubiquitination. Specifically, radiation-induced ATM-dependent phosphorylation of DGCR8 at serine 677 facilitates USP51 to bind, deubiquitinate, and stabilize DGCR8, which leads to the recruitment of DGCR8 and DGCR8’s binding partner RNF168 to MDC1 and RNF8 at DSBs. This, in turn, promotes ubiquitination of histone H2A, repair of DSBs, and radioresistance. Altogether, these findings reveal the non-canonical function of DGCR8 in DSB repair and suggest that radiation treatment may result in therapy-induced tumor radioresistance through ATM- and USP51-mediated activation and upregulation of DGCR8.
Highlights
In response to DNA double-strand breaks (DSBs), repair proteins are recruited to the damaged sites
By performing Co-IP experiments to examine the interactions between endogenous proteins, we found that knockdown of DGCR8 in LM2 cells blocked ionizing radiation (IR)-induced RNF168RNF8 and RNF168-MDC1 interactions, without inhibiting the RNF8-MDC1 interaction (Fig. 3d), which suggests that DGCR8 is required for recruiting RNF168 to MDC1 and RNF8 after irradiation
Similar interaction patterns were observed with or without Micrococcal Nuclease (MNase) treatment (Fig. 3e). These results indicate that the interactions of DGCR8 with RNF168, RNF8, and MDC1 occur on chromatin, but these protein–protein interactions are not mediated by chromatin
Summary
In response to DNA double-strand breaks (DSBs), repair proteins are recruited to the damaged sites. Radiation-induced ATM-dependent phosphorylation of DGCR8 at serine 677 facilitates USP51 to bind, deubiquitinate, and stabilize DGCR8, which leads to the recruitment of DGCR8 and DGCR8’s binding partner RNF168 to MDC1 and RNF8 at DSBs. This, in turn, promotes ubiquitination of histone H2A, repair of DSBs, and radioresistance. Dicer and Drosha can generate dsRNAs that promote DNA repair by facilitating the recruitment of DDR factors[41,42,43] or through the formation of DNA:RNA hybrids around DNA break sites[44] Both Dicer and DGCR8 have been reported to regulate nucleotide excision repair of ultraviolet-induced lesions (pyrimidine dimers and base modifications)[45,46]. DGCR8 and its constitutive binding partner RNF168 in turn interact with MDC1 and RNF8 to promote histone ubiquitination, DSB repair, and radioresistance
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