DNA polymerase η promotes nonhomologous end joining upon etoposide exposure dependent on the scaffolding protein Kap1

Ma, Xinwen; Wang, Chen; Zhou, Bo; Cheng, Zina; Mao, Zhiyong; Tang, Tie-Shan; Guo, Caixia

doi:10.1016/j.jbc.2022.101861

Cited by 4 publications

(3 citation statements)

References 44 publications

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“…Human U2OS and HEK293T cells were obtained from the American Type Culture Collection (Rockville, MD). RAD18 knockout (RAD18-/-) cells were established using TALEN as described previously [ 5 ]. These cell lines were grown in DMEM medium supplemented with 10% fetal bovine serum.…”

Section: Methodsmentioning

confidence: 99%

“…The whole cell lysates were immunoprecipitated with either anti-Flag M2 agarose or GFP nanobody agarose beads. RAD18 O-GlcNAcylation was confirmed by denatured immunoprecipitation (IP) as previously described [ 5 , 30 ]. Briefly, cells were lysed with 1xSDS lysis buffer (50 mM Tris-HCl pH 6.8, 100 mM DTT, 2% SDS, 10% glycerol) at 95 °C for 15 min.…”

Section: Methodsmentioning

confidence: 99%

“…The E3 ubiquitin ligase RAD18 coordinates translesion DNA synthesis (TLS), homologous recombination (HR) and other DNA damage response pathways to maintain genome integrity [1][2][3][4][5][6]. As a conserved and predominant DNA damage tolerance mechanism, TLS is regulated by the monoubiquitination of proliferating cell nuclear antigen (PCNA), which acts as a ringshaped homotrimer encircling DNA as a scaffold to promote multiple specialized TLS polymerases to replicative bypass ultraviolet (UV)-and cisplatin (cis-diamminedichloroplatinum, CDDP)-induced damage [6][7][8].…”

Section: Introductionmentioning

confidence: 99%

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RAD18 O-GlcNAcylation promotes translesion DNA synthesis and homologous recombination repair

Ma,

Fu,

Sun

et al. 2024

Cell Death Dis

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RAD18, an important ubiquitin E3 ligase, plays a dual role in translesion DNA synthesis (TLS) and homologous recombination (HR) repair. However, whether and how the regulatory mechanism of O-linked N-acetylglucosamine (O-GlcNAc) modification governing RAD18 and its function during these processes remains unknown. Here, we report that human RAD18, can undergo O-GlcNAcylation at Ser130/Ser164/Thr468, which is important for optimal RAD18 accumulation at DNA damage sites. Mechanistically, abrogation of RAD18 O-GlcNAcylation limits CDC7-dependent RAD18 Ser434 phosphorylation, which in turn significantly reduces damage-induced PCNA monoubiquitination, impairs Polη focus formation and enhances UV sensitivity. Moreover, the ubiquitin and RAD51C binding ability of RAD18 at DNA double-strand breaks (DSBs) is O-GlcNAcylation-dependent. O-GlcNAcylated RAD18 promotes the binding of RAD51 to damaged DNA during HR and decreases CPT hypersensitivity. Our findings demonstrate a novel role of RAD18 O-GlcNAcylation in TLS and HR regulation, establishing a new rationale to improve chemotherapeutic treatment.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%