Tax cooperates with p300 to induce histone hyperacetylation at late origins. (A) HeLa cells were transfected with different Tax expression vectors (wild-type Tax, M22, M47, K88A, and Δ2-22) and synchronized at G₁/S transition with HU. Ctrl corresponds to cells transfected with an empty vector. BrdU incorporation was performed as described in Figure 2D. (B) JPX9 and JPX9i cells were treated with CSK buffer and then fixed and analyzed by confocal microscopy using indicated antibodies. Fluorescence intensity of cells in G₁ was measured as described in supplemental Figure 8. (C) Rat-1 cells were transfected with expression vectors for wild-type Tax or K88A mutant and synchronized at G₁/S transition with HU before CSK extraction, fixation, and confocal microscopy analysis using α-H3K56ac and α-Tax antibodies (see supplemental Figure 9). Mean fluorescence intensity of H3K56ac labeling was analyzed in control (Ctrl), wild-type Tax, or mutant K88A-positive cells (n = 30). (D) p300, H3K9-14ac, and H3K56ac occupancy at Myc, LaminB2, β-globin, and RNF185 replication origins was detected by ChIP in JPX9 cells. ChIP results were expressed as a percentage of input DNA. Representative results of 1 of 3 independent ChIP experiments are shown. (E) Replication timing analysis at Myc, LaminB2, β-globin, and RNF185 replication origins was determined by isolation of nascent DNA in JPX9 (■) or JPX9i cells (□) sorted in G₁, S_1-4, and G₂ stages of the cell cycle. NS indicates not significant. **P < .01, ***P < .001, according to Kruskal-Wallis test.