Fig. 1. Mitotic repression of pol III transcription is mediated by phosphorylation. (A) Class III genes are transcribed more efficiently by asynchronous extracts than by mitotic extracts. pHu5S3.1 (lanes 1 and 2) or pVA1 (lanes 3 and 4) templates (500 ng) were transcribed using 14 µg of extract prepared from either asynchronous (lanes 1 and 3) or mitotic (lanes 2 and 4) cells. (B) Mitotic repression of pol III transcription can be relieved using a kinase inhibitor and accentuated with a phosphatase inhibitor. pGlu6 (500 ng) was transcribed using 30 µg of either asynchronous (lanes 1–3) or mitotic extract (lanes 4–6). Reactions in lanes 1 and 4 contained 2.4 mM DMAP; those in lanes 2 and 5 contained 2 µM okadaic acid.