Functional association of Gdown1 with RNA polymerase II poised on human genes.

Most human genes are loaded with promoter-proximally paused RNA polymerase II (Pol II) molecules that are poised for release into productive elongation by P-TEFb. We present evidence that Gdown1, the product of the POLR2M gene that renders Pol II responsive to Mediator, is involved in Pol II elongation control. During in vitro transcription, Gdown1 specifically blocked elongation stimulation by TFIIF, inhibited the termination activity of TTF2, and influenced pausing factors NELF and DSIF, but did not affect the function of TFIIS or the mRNA capping enzyme.

Functional coupling of cleavage and polyadenylation with transcription of mRNA.

Cleavage and polyadenylation define the 3' ends of almost all eukaryotic mRNAs and are thought to occur during transcription. We describe a human in vitro system utilizing an immobilized template, in which transcripts in RNA polymerase II elongation complexes are efficiently cleaved and polyadenylated. Because the cleavage rate of free RNA is much slower, we conclude that cleavage is functionally coupled to transcription.

Binding of the 7SK snRNA turns the HEXIM1 protein into a P-TEFb (CDK9/cyclin T) inhibitor.

The positive transcription elongation factor b (P-TEFb) plays a pivotal role in productive elongation of nascent RNA molecules by RNA polymerase II. Core active P-TEFb is composed of CDK9 and cyclin T. In addition, mammalian cell extracts contain an inactive P-TEFb complex composed of four components, CDK9, cyclin T, the 7SK snRNA and the MAQ1/HEXIM1 protein.

Cotranscriptional processing of Drosophila histone mRNAs.

The 3' ends of metazoan histone mRNAs are generated by specialized processing machinery that cleaves downstream of a conserved stem-loop structure. To examine how this reaction might be influenced by transcription, we used a Drosophila melanogaster in vitro system that supports both processes. In this system the complete synthesis of histone mRNA, including transcription initiation and elongation, followed by 3' end formation, occurred at a physiologically significant rate.