The binding of the alpha subunit of protein kinase CK2 and RAP74 subunit of TFIIF to protein-coding genes in living cells is DRB sensitive.

In a previous report, we documented that a major portion of the nuclear protein kinase CK2alpha (CK2alpha) subunit does not form heterooligomeric structures with the beta subunit, but it binds tightly to nuclear structures in an epithelial Chironomus cell line. We report here that the CK2alpha, but not beta, subunit is co-localized with productively transcribing RNA polymerase II (pol II) on polytene chromosomes of Chironomus salivary gland cells. Likewise, the RAP74 subunit ofTFIIF, a potential substrate for CK2, is co-localized with pol II.

Heat-shock-specific phosphorylation and transcriptional activity of RNA polymerase II.

The carboxyl-terminal domain (CTD) of the largest RNA polymerase II (pol II) subunit is a target for extensive phosphorylation in vivo. Using in vitro kinase assays it was found that several different protein kinases can phosphorylate the CTD including the transcription factor IIH-associated CDK-activating CDK7 kinase (R. Roy, J.

Phosphorylation dependence of the initiation of productive transcription of Balbiani ring 2 genes in living cells.

Using polytene chromosomes of salivary gland cells of Chironomus tentans, phosphorylation state-sensitive antibodies and the transcription and protein kinase inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB), we have visualized the chromosomal distribution of RNA polymerase II (pol II) with hypophosphorylated (pol IIA) and hyperphosphorylated (pol II0) carboxyl-terminal repeat domain (CTD). DRB blocks labeling of the CTD with 32Pi within minutes of its addition, and nuclear pol II0 is gradually converted to IIA; this conversion parallels the reduction in transcription of protein-coding genes. DRB also alters the chromosomal distribution of II0: there is a time-dependent clearance from chromosomes of phosphoCTD (PCTD) after addition of DRB, which coincides in time with the completion and release of preinitiated transcripts.

The salivary gland 42-kDa phosphoprotein is a single-stranded DNA-binding protein with characteristics of the epithelial casein kinase N42 in Chironomus tentans.

The DNA-binding and phosphorylation properties of a rapidly phosphorylated nuclear 42-kDa phosphoprotein and of its two structurally related proteins, pp43 and pp44 in Chironomus tentans salivary glands were investigated. pp42, pp43 and pp44 bind promoter probes of the ecdysterone controlled I-18C gene and of the joint histone H2A/H2B genes in a sequence-selective and single-stranded DNA (ssDNA) specific manner. Rapid phosphorylation appears to give pp42 and pp43 uniquely hydrophilic characters making them soluble in the aqueous phase during phenol treatment.

Analysis of a novel DNA-binding protein kinase CKII-like enzyme of Chironomus cells.

We have previously described a Chironomus tentans nuclear 42 kDa phosphoprotein preferentially associated with transcriptionally active chromatin. In an attempt to purify and identify the kinase responsible for the phosphorylation of the 42 kDa protein, a casein-phosvitin affinity chromatography was used. Unexpectedly, in the eluted kinase fraction, a novel 42 kDa casein kinase, designated protein kinase CK42, with a kinase activity similar, but not identical, to protein kinase CKII, could be identified.