Figure 8. ZRF1 regulates XPC ubiquitylation. (A) ZRF1 facilitates XPC ubiquitylation after UV irradiation. Whole-cell extracts of control and ZRF1 knockdown HEK293T cells from the stated time points were subjected to Western blotting and probed with the indicated antibodies. (B) Role of RING1B and ZRF1 in XPC ubiquitylation. Control cells and RING1B and ZRF1 knockdown HEK293T cells expressing HAUbiquitin were irradiated with UV light. After immunoprecipitation with HA-specific antibody, the precipitated material was subjected to Western blotting and blots were incubated with the indicated antibodies. Inputs correspond to 5%. (C) Control cells and RING1B and ZRF1 knockdown HEK293T cells expressing HAXPC and HISUbiquitin were irradiated with UV light. After immunoprecipitation with HA-specific antibody, the precipitated material was subjected to Western blotting and blots were incubated with the indicated antibodies. Inputs correspond to 5%. (D) Control cells and RING1B and ZRF1 knockdown HEK293T cells expressing HISUbiquitin were irradiated with UV and harvested 1 h after UV exposure. Ubiquitylated proteins were purified by NiNTA agarose under denaturing conditions, and Western blots of the purified material were incubated with the indicated antibodies. (E) The UV–RING1B complex and ZRF1 cooperate during NER. DNA lesions (yellow star) are recognized by the UV-RING1B complex (DDB1–DDB2–CUL4B–RING1B), which catalyzes ubiquitylation of histone H2A (gray sphere). ZRF1 is recruited to the lesion site by XPC and tethers to the H2A-ubiquitin mark. ZRF1 causes the assembly of the UV–DDB–CUL4A complex, which subsequently catalyzes ubiquitylation of XPC.
Figure 8.

ZRF1 regulates XPC ubiquitylation. (A) ZRF1 facilitates XPC ubiquitylation after UV irradiation. Whole-cell extracts of control and ZRF1 knockdown HEK293T cells from the stated time points were subjected to Western blotting and probed with the indicated antibodies. (B) Role of RING1B and ZRF1 in XPC ubiquitylation. Control cells and RING1B and ZRF1 knockdown HEK293T cells expressing HAUbiquitin were irradiated with UV light. After immunoprecipitation with HA-specific antibody, the precipitated material was subjected to Western blotting and blots were incubated with the indicated antibodies. Inputs correspond to 5%. (C) Control cells and RING1B and ZRF1 knockdown HEK293T cells expressing HAXPC and HISUbiquitin were irradiated with UV light. After immunoprecipitation with HA-specific antibody, the precipitated material was subjected to Western blotting and blots were incubated with the indicated antibodies. Inputs correspond to 5%. (D) Control cells and RING1B and ZRF1 knockdown HEK293T cells expressing HISUbiquitin were irradiated with UV and harvested 1 h after UV exposure. Ubiquitylated proteins were purified by NiNTA agarose under denaturing conditions, and Western blots of the purified material were incubated with the indicated antibodies. (E) The UV–RING1B complex and ZRF1 cooperate during NER. DNA lesions (yellow star) are recognized by the UV-RING1B complex (DDB1–DDB2–CUL4B–RING1B), which catalyzes ubiquitylation of histone H2A (gray sphere). ZRF1 is recruited to the lesion site by XPC and tethers to the H2A-ubiquitin mark. ZRF1 causes the assembly of the UV–DDB–CUL4A complex, which subsequently catalyzes ubiquitylation of XPC.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal