Figure 2.

YY1 DNA-binding activity is lost in prometaphase. WCEs were prepared from asynchronous (Asy), nocodazole-arrested (Noc), or thymidine-arrested HeLa cells (Th), separated by SDS-PAGE, and analyzed by Western blot using anti-YY1 (H-10), anti-cyclin B, or anti-GAPDH antibodies (A) and electrophoretic mobility shift assay (EMSA) using ³²P-labeled H3.2α as probe (B). The specificity of the YY1 shift was tested by addition of anti-YY1 (C-20), anti-GFP, or anti-Sp1 to the WCE from asynchronous cells as indicated. The cell cycle distribution of the cells was assessed by FACS analysis (C). Confocal images of nocodazole-arrested cells are presented in Supplementary Figure S2.