Casein kinase type II is involved in the inhibition by 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole of specific RNA polymerase II transcription.

Abstract

We have described a HeLa protein kinase whose activity is inhibited by the nucleotide analogue 5,6dichloro-1-8-D-ribofuranosylbenzimidazole (DRB) at concentrations similar to those required to inhibit in vivo and in vitro specific transcription (Zandomeni, R., and Weinmann, R. (1984) J. Biol. Chem. 259, 14804-14822). We have now detected an analogous DRB-sensitive kinase from calf thymus and purified it to homogeneity. Based on the subunit composition of the enzyme and other common biochemical and chromatographic properties, we identified it as casein kinase 11. The extent of DRB inhibition of the purified calf thymus enzyme is indistinguishable from that observed for inhibition of in vitro transcription with the HeLa cell extract. The DRB bromo-derivative, 5,6dibromo-1-8-D-ribofuranosylbenzimidazole is a more potent inhibitor of in vivo transcription and inhibits purified casein kinase I1 activity and specific in vitro transcription at 6-10 times lower concentrations than DRB. Moreover, addition of an excess of the purified calf thymus casein kinase I1 enzyme to a HeLa in vitro transcription reaction inhibited by DRB partially overcomes this inhibition. Thus, we conclude that casein kinase I1 is involved directly or indirectly in the inhibition by DRB of specific RNA polymerase 11-mediated transcription. This demonstrates the participation of a protein kinase in a eukaryotic RNA polymerase 11-specific transcription system. The adenosine analogue 5,6-dichloro-l-~-~-ribofuranosylbenzimidazole (DRB1) is widely used to inhibit mRNA synthesis (l), acting rapidly on mRNA precursor synthesis both in mammalian cells in culture and in insects. Using i n vitro RNA polymerase I1 transcription systems that reproduce faithful i n uiuo initiation conditions, we have shown that the concentrations of DRB required for inhibition are similar to those effective in vivo (2) and that RNA elongation of prein-