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The RNA polymerase II CTD is essential for 3' end cleavage of metazoan pre-mRNAs and binds 3' end processing factors in vitro. We show genetic and biochemical interactions between the CTD and the Pcf11 subunit of the yeast cleavage/polyadenylation factor, CFIA. In vitro binding to Pcf11 required phosphorylation of the CTD on Ser2 in the YSPTSPS heptad(More)
The universal pre-mRNA processing events of 5' end capping, splicing, and 3' end formation by cleavage/polyadenylation occur co-transcriptionally. As a result, the substrate for mRNA processing factors is a nascent RNA chain that is being extruded from the RNA polymerase II exit channel at 10-30 bases per second. How do processing factors find their(More)
Messenger RNA is produced by RNA polymerase II (pol II) transcription, followed by processing of the primary transcript. Transcription, splicing and cleavage-polyadenylation can occur independently in vitro, but we demonstrate here that these processes are intimately linked in vivo. We show that the carboxy-terminal domain (CTD) of the pol II large subunit(More)
Changes in promoter structure and occupation have been shown to modify the splicing pattern of several genes, evidencing a coupling between transcription and alternative splicing. It has been proposed that the promoter effect involves modulation of RNA pol II elongation rates. The C4 point mutation of the Drosophila pol II largest subunit confers on the(More)
The C-terminal heptad repeat domain (CTD) of RNA polymerase II (pol II) is proposed to target pre-mRNA processing enzymes to nascent pol II transcripts, but this idea has not been directly tested in vivo. In vitro, the yeast mRNA capping enzymes Ceg1 and Abd1 bind specifically to the phosphorylated CTD. Here we show that yeast capping enzymes cross-link in(More)
We have investigated the role of the RNA Polymerase II (Pol II) carboxy-terminal domain (CTD) in mRNA 5' capping. Transcripts made in vivo by Pol II with a truncated CTD had a lower proportion of capped 5' ends than those made by Pol II with a full-length CTD. In addition, the enzymes responsible for cap synthesis, RNA guanylyltransferase, and RNA(More)
Much of the complex process of RNP biogenesis takes place at the gene cotranscriptionally. The target for RNA binding and processing factors is, therefore, not a solitary RNA molecule but, rather, a transcription elongation complex (TEC) comprising the growing nascent RNA and RNA polymerase traversing a chromatin template with associated passenger proteins.(More)
Maturation of mRNA precursors often occurs simultaneously with their synthesis by RNA polymerase II (Pol II). The co-transcriptional nature of mRNA processing has permitted the evolution of coupling mechanisms that coordinate transcription with mRNA capping, splicing, editing and 3' end formation. Recent experiments using sophisticated new methods for(More)
Promoter and enhancer elements can influence alternative splicing, but the basis for this phenomenon is not well understood. Here we investigated how different transcriptional activators affect the decision between inclusion and exclusion (skipping) of the fibronectin EDI exon. A mutant of the acidic VP16 activation domain called SW6 that preferentially(More)
Processing of RNA precursors to their mature form often occurs co-transcriptionally. Consequently, the ternary complex of DNA template, RNA polymerase and nascent RNA chain is the physiological substrate for factors that modify the nascent RNA by capping, splicing and cleavage/polyadenylation. mRNA production is thought to occur within a "factory" that(More)