Figure 1.

Mitotic repression of transcription is abolished by roscovitine treatment. HeLa cells blocked in mitosis by colchicine treatment (0.02 μg/ml for 14 h) were treated (d–f) or not (a–c) with 150 μM roscovitine, a specific inhibitor of cdc2–cyclin B kinase, for 30 min. Mitotic cells were also treated with 0.5 μM okadaic acid for 1 h before addition of 150 μM roscovitine for 30 min (g–i). Transcription activity was revealed by the detection of in situ incorporation of BrUTP in conditions favoring both RNA pol I and RNA pol II activity (b, e, and h). Simultaneously, the inhibition of cdc2–cyclin B kinase was verified by the detection of the hyperphosphorylated form of histone H1 (c, f, and i). The chromosomes were stained with DAPI (a, d, and g). In control cells (a–c) no transcription activity was detected (b) and histone H1 was hyperphosphorylated (c). In roscovitine-treated cells (d–f) transcription activity was detected (e) in mitotic cells negative for hyperphosphorylated histone H1 (f). Arrowheads indicate cells positive for phosphorylation of histone H1 (f) and negative for transcription activity (e). In mitotic cells, additionally pretreated with okadaic acid (g–i), neither dephosphorylation of histone H1 (i) nor transcription activity (h) was detected. The inhibition of cdc2–cyclin B kinase abolished mitotic repression of transcription dependent on an okadaic acid–sensitive phosphatase. Bar, 10 μm.