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J. Biol. Chem., Vol. 278, Issue 9, 7180-7188, February 28, 2003
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From the RNA polymerase II (pol II) is subject to an early
elongation delay induced by negative factors Spt5/Spt4 and NELF,
which is overcome by the positive factor P-TEFb (Cdk9/cyclin T), a
protein kinase that phosphorylates the pol II C-terminal domain (CTD) and the transcription elongation factor Spt5. Although the
rationale for this arrest and restart is unclear, recent studies
suggest a connection to mRNA capping, which is coupled to
transcription elongation via physical and functional interactions
between the cap-forming enzymes, the CTD-PO4, and
Spt5. Here we identify a novel interaction between fission yeast RNA
triphosphatase Pct1, the enzyme that initiates cap formation, and
Schizosaccharomyces pombe Cdk9. The C-terminal segment of
SpCdk9 comprises a Pct1-binding domain distinct from the N-terminal Cdk
domain. We show that the Cdk domain interacts with S. pombe
Pch1, a homolog of cyclin T, and that the purified recombinant
SpCdk9/Pch1 heterodimer can phosphorylate both the pol II CTD and the
C-terminal domain of S. pombe Spt5. We provide genetic
evidence that SpCdk9 and Pch1 are functional orthologs of the
Saccharomyces cerevisiae CTD kinase Bur1/Bur2, a putative
yeast P-TEFb. Mutations of the kinase active site and the regulatory
T-loop of SpCdk9 abolish its activity in vivo. Deleting the
C-terminal domain of SpCdk9 causes a severe growth defect. We suggest a
model whereby Spt5-induced arrest of early elongation ensures a
temporal window for recruitment of the capping enzymes, which in turn
attract Cdk9 to alleviate the arrest. This elongation checkpoint
may avoid wasteful rounds of transcription of uncapped
pre-mRNAs.
Interactions between Fission Yeast Cdk9, Its Cyclin Partner
Pch1, and mRNA Capping Enzyme Pct1 Suggest an Elongation Checkpoint
for mRNA Quality Control*
,¶
Molecular Biology Program, Sloan-Kettering
Institute, New York, New York 10021 and the § Department
of Microbiology and Immunology, Weill Medical College of Cornell
University, New York, New York 10021
*
This work was supported by National Institutes of Health
Grant GM52470.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Copyright © 2003 by The American Society for Biochemistry and Molecular Biology, Inc.
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