Fig. 4: UVSSA stabilizes the CSA–ELOF1 interface and stimulates Pol II ubiquitylation.
a, Zoom-in on the interfaces among UVSSA, ELOF1 and CSA. b, TCR-dependent γH2AX induction after trabectedin exposure in replicating cells labeled with 5-EdU in the indicated RPE1-iCas9 cells. The γH2AX levels were normalized to trabectedin-treated WT cells within each experiment. The experiment was performed three times. Each colored circle represents one cell. Each black circle represents the mean of two technical replicates, with more than 70 cells collected per technical replicate. The black lines represent the mean of all three independent experiments. c, Transcription recovery (RRS) by 5-EU labeling in the indicated RPE1-iCas9 cells after UV irradiation (24 h; 9 J m−2). The 5-EU levels were normalized to mock treatment for each cell line. The experiment was performed three times. Each black circle represents the mean of two technical replicates. The black lines represent the mean of all three independent experiments. d, Percentage of particles with CSA docked onto ELOF1 in the Pol II–CSB–CSA–ELOF1 dataset (left) and the Pol II–CSB–CSA–ELOF1-UVSSA dataset (right). e, Endogenous Pol II-S2 immunoprecipitation (IP) after UV irradiation (9 J m−2, 1-h recovery) in RPE1-iCas9-WT or UVSSA-KO cells complemented with GFP-tagged UVSSA. TCR complex assembly and Pol II ubiquitylation were analyzed with the indicated antibodies. The data shown represent at least three independent experiments. f, In vitro ubiquitylation assay of Pol II in the presence of CSB, CRL4CSA and ELOF1 with increasing amounts of UVSSA. The experiment was repeated two times.
