Fig. 3: RTF2 regulates RNase H2 localization to the replisome.
From: RTF2 controls replication repriming and ribonucleotide excision at the replisome
a Schematic of the iPOND set-up in SV40-immortlized Rtf2-/- MEFs expressing HA-FLAG empty vector (EV) or RTF2 cDNA. EdU pulses were normalized to replication tract lengths. Nascent DNA was purified using streptavidin beads against biotin-conjugated EdU. b Peptide Spectral Match (#PSM) values for indicated proteins from experiment in (a). c Representative iPOND immunoblot from p53-/- MEFs at 72 hr after Cre. d Left: Representative images of RNASEH2A-EdU nascent proximity ligation assay (nPLA) co-stained with PCNA in SV40-immortalized Rtf2-/- MEFs expressing HA-FLAG EV or RTF2 cDNA. Right: Quantification of RNASEH2A-EdU foci in PCNA-positive cells. e Representative iPOND immunoblot in DDI-depleted U2OS cells. Proteins purified using streptavidin beads recognizing nascent DNA were detected by immunoblot. f Representative iPOND immunoblot in CRISPR-edited RNASEH2A KO or WT HeLa cells. Proteins purified using streptavidin beads recognizing nascent DNA were detected by immunoblot. Experiments were conducted at least three times in biological replicates with consistent results for (c)–(f). Mean is shown with red line for (d). Significance was evaluated by Kruskal-Wallis ANOVA with a Dunn’s post-test. P = Parental, RH2A = RNASEH2A, RH2B = RNASEH2B, RH2C = RNASEH2C, RH2 = RNASEH2 complex, Ctrl = Control. Source data are provided as a Source Data file.
