Fig. 1: Dual mono-ubiquitylated PAF15 (PAF15Ub2) specifically binds to replicating chromatin.
From: Two distinct modes of DNMT1 recruitment ensure stable maintenance DNA methylation
a xDNMT1 pull-downs from native chromatin extracts were analyzed by LC-MS/MS. The volcano plot summarizes the quantitative results and highlights the interacting proteins enriched upon addition of ubiquitin to UbVS-treated extracts. b Xenopus interphase egg extracts were added with sperm chromatin and incubated in the absence or presence of His6-ubiquitin (58 μM final). Chromatin-bound proteins were isolated and analyzed by immunoblotting using the indicated antibodies. For PAF15 levels in the extracts, see Supplementary Fig. 1f. c Interphase egg extracts were added with sperm chromatin and incubated in the presence of 15 μM aphidicolin (Aph) or in its absence (DMSO). Chromatin-bound proteins were isolated and analyzed by immunoblotting using the indicated antibodies. d PAF15-deleted extracts were supplemented with wild-type xPAF15-Flag3 and its variants (K18R, K27R, and K18R/K27R). After the addition of sperm chromatin, chromatin-bound proteins were isolated and analyzed by immunoblotting using the indicated antibodies. The extracts were also analyzed by immunoblotting. e PAF15-depleted extracts were supplemented with wild-type xPAF15-Flag3, its PIP-box mutant (FF/AA), or K18R/K27R mutant (KRKR). After the addition of sperm chromatin, chromatin-bound proteins were isolated and analyzed by immunoblotting using the indicated antibodies. The extracts were also analyzed by immunoblotting. Source data are provided as a Source Data file.
