The ubiquitin-selective segregase VCP/p97 orchestrates the response to DNA double-strand breaks

Abstract

Unrepaired DNA double-strand breaks (DSBs) cause genetic instability that leads to malignant transformation or cell death. Cells respond to DSBs with the ordered recruitment of signalling and repair proteins to the site of lesion. Protein modification with ubiquitin is crucial for the signalling cascade, but how ubiquitylation coordinates the dynamic assembly of these complexes is poorly understood. Here, we show that the human ubiquitin-selective protein segregase p97 (also known as VCP; valosin-containing protein) cooperates with the ubiquitin ligase RNF8 to orchestrate assembly of signalling complexes and efficient DSB repair after exposure to ionizing radiation. p97 is recruited to DNA lesions by its ubiquitin adaptor UFD1-NPL4 and Lys-48-linked ubiquitin (K48-Ub) chains, whose formation is regulated by RNF8. p97 subsequently removes K48-Ub conjugates from sites of DNA damage to orchestrate proper association of 53BP1, BRCA1 and RAD51, three factors critical for DNA repair and genome surveillance mechanisms. Impairment of p97 activity decreases the level of DSB repair and cell survival after exposure to ionizing radiation. These findings identify the p97-UFD1-NPL4 complex as an essential factor in ubiquitin-governed DNA-damage response, highlighting its importance in guarding genome stability.

Proliferating Cell Nuclear Antigen (PCNA)-binding Protein C1orf124 Is a Regulator of Translesion Synthesis

“…VCP/p97 has also been shown to function in repair of ionizing radiationinduced double-strand breaks. VCP/p97 is recruited to ionizing radiation-induced damage sites in an RNF8-dependent manner, where it catalyzes the removal of the Lys-48-conjugated protein substrates to allow for proper assembly of downstream signaling effectors, including RAD51, BRCA1, and 53BP1 (37). In addition, VCP/p97 also mediates the removal of the RNA polymerase II complex when it is stalled at UV light-induced DNA lesions (38).…”

K48-linked ubiquitination and protein degradation regulate 53BP1 recruitment at DNA damage sites

“…Hence, the local enrichment of H4K20(me2) triggered by MMSET does not explain the requirement of RNF8 for the recruitment of 53BP1 [5][6][7]. It was initially thought that RNF8 only catalyzes K63-linked ubiquitination, but multiple recent reports describe the ability of RNF8 to stimulate the formation of K48-linked ubiquitin chains [4,14,15], confirming a role for RNF8-mediated degradation or chromatin extraction during the DNA damage response.…”

“…Furthermore, depletion of the proteasomal subunits PSMB3 and PSMD4 inhibits the formation of FANCD2 foci following ionizing radiation [3]. In addition, K48-linked polyubiquitin chains accumulate at sites of DNA damage, suggesting a role for protein degradation during the DNA damage response [4]. However, the molecular mechanisms controlling protein ubiquitination and degradation following DNA damage are still unclear.…”

“…VCP/p97 is recruited to DNA breaks in an RNF8-dependent fashion requiring the formation of K48-linked ubiquitin chains [4]. The presence of VCP/p97 at DNA lesions is a prerequisite for 53BP1 foci formation and may promote the chromatin extraction of ubiquitinated proteins Figure 1 K48-linked ubiquitination regulates the recruitment of 53BP1 to DNA lesions.…”

“…This enables the local recruitment of 53BP1 to DNA damage sites. npg [4]. In fact, the Polycomb group protein L3MBTL1 is removed from DNA lesions by an RNF8-and VCP/p97-dependent mechanism [16].…”