TopI binds RNAP II and promotes RNAP II elongation on α-satellite chromatin a TopI inhibition decreases RNAP II-pSer2 levels on α-satellite DNAs. HeLa cells treated with DMSO or CPT (2.5 µM) for 12 h were subjected to chromatin immunoprecipitation analysis with anti-RNAP II-pSer2 antibody. Biological replicates (n = 1 for beads, n = 4 for DMSO and CPT). b TopI inhibition reduces RNAP II-pSer2 levels on α-satellite DNAs in G1 cells. Stretched chromatin was prepared from thymidine-arrested HeLa cells treated with DMSO or CPT (2.5 µM) for 6 h and immunostaining was performed. n = 3 biological replicates. c Mitosis-specific inhibition of TopI reduces RNAP II-pSer2 levels on α-satellite DNAs. Mitotic HeLa cells were enriched by a brief treatment of nocodazole (5 µM). Collected mitotic cells were further treated with DMSO or CPT (2.5 µM) for 5 h and then subjected for chromosome spread followed by immunostaining. n = 3 biological replicates. d TopI physically interacts with RNAP II and phospho-RNAP II (pSer2). Lysates of HeLa cells transfected Luciferase or Top1 siRNAs (#1) were incubated with IgG or antibody against TopI. Pelleted proteins were blotted with the indicated antibodies. The physical interaction between TopI and non-phospho-RNAP II was recorded in Supplementary fig. 1e. “H” high exposure. e TopI and TopI-cleavage complex (cc) are present on the centromere in interphase. Stretched chromatin was prepared from log-phase HeLa cells and immunostaining was performed. Similar results were observed in at least two biological replicates. f TopI-cc is enriched on the centromere in mitosis. HeLa cells were briefly treated with nocodazole (5 µM) and mitotic cells were collected for chromosome spread followed by immunostaining. Similar results were observed in at least two biological replicates. Validation of TopI and Top-cc fluorescence signals were recorded in Supplementary Figs. 1e, f. All data here are presented as mean values +/− SEM. Two-sided Student’s T-test for (a–c). ns, not significant (P > 0.1). Numeric values for P < 0.1 are shown. The source data are provided as Source Data file.