The transition between transcription initiation and elongation requires a significant exchange of the factors associated with RNApII. The basal transcription factors and Mediator at the promoter must release the elongating RNApII. Elongation factors, such as the Spt4/5, TREX, and Paf complexes, subsequently associate with RNApII throughout the transcribed region (Kaplan et al, 2000
; Pokholok et al, 2002
). Although serine 2 phosphorylation of the Rpb1 CTD mediates the coupling of transcription with polyadenylation at 3′-ends (Ahn et al, 2004
; Ni et al, 2004
), our data suggest that Ctk1 also affects the transition from transcription initiation to elongation at 5′-ends of genes by promoting dissociation of basal factors from polymerase.
Surprisingly, this 5′ function of Ctk1 does not require kinase activity (). Thus, it is most likely that Ctk1 has a structural function, either directly displacing basal factors or by recruiting elongation factors to the polymerase (perhaps through protein–protein contacts independent of phosphorylation) that subsequently mediate the dissociation of basal transcription factors from elongating RNApII. One possibility is that the Ctk1/Ctk2 and Kin28/Ccl1 CTD kinase/cyclin modules occupy the same physical space near the Rpb1 CTD, making their binding mutually exclusive. In the absence of Ctk1, contacts between RNApII and basal factors apparently persist during transcription elongation, resulting in the downstream cross-linking of basal factors ().
After initiation of transcription in vitro
, the basal transcription factors TFIID, TFIIA, TFIIH, and TFIIE, as well as the activator and Mediator, remain at the promoter in the scaffold complex. This complex may facilitate subsequent rounds of transcription reinitiation (Yudkovsky et al, 2000
; Hahn, 2004
). Hahn and colleagues observed that the kinase activities of the TFIIH subunit Kin28 and the Mediator Subunit Srb10 contribute to the dissociation of polymerase, TFIIB, and TFIIF from scaffolds (Yudkovsky et al, 2000
; Liu et al, 2004
). Our results suggest that Ctk1 contributes to scaffold maintenance. In ctk1
Δ extracts, both scaffold and non-scaffold factors dissociate from template DNA after a single round of transcription (). This result is consistent with a model where Ctk1 promotes the release of basal transcription factors from transcribing RNApII as it enters productive elongation.
There are at least two possible variations of this model that could explain the ChIP results (). In the first, basal factors release from the promoter in ctk1Δ cells, but remain associated with the polymerase as it moves through the gene body. In the second, RNApII actually remains tethered to the basal factors at the promoter while transcribing. This would result in the transcribing polymerase ‘pulling' the transcribed region past basal factors, thereby allowing cross-linking with downstream sequences.
Although a function for Ctk1 in the 5′ transcriptional transition is a new finding, the presumed mammalian homologue Cdk9/P-TEFb has been shown previously to modulate early elongation (Chao and Price, 2001
; Yamada et al, 2006
). At least part of Cdk9 function at 5′-ends is mediated by its kinase activity, but it is certainly possible that there are additional structural functions. There are several steps before productive transcription elongation, including promoter proximal pausing, clearance, and escape (Goodrich and Tjian, 1994
); further work will be required to determine exactly how Ctk1 and Cdk9 affect each of these.