Ssu72, a component of the CPF 3'-processing machinery in yeast, is an RNAPII CTD phosphatase with specificity for serine-5.
Krishnamurthy Shankarling (1), Xiaoyuan He (2), Mariela Reyes-Reyes (1), Claire Moore (2), Michael Hampsey (1)
(1) Department of Biochemistry, RWJ Medical School, UMDNJ, 683, Hoes Lane West, Piscataway, NJ, 08854, USA;
(2) Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, MA 02111
The carboxy-terminal domain CTD of the largest subunit of RNA polymerase II (RNAP II) is composed of a heptapeptide (YSPTSPS) repeat whose sequence is reiterated 26 times in S. cerevisiae. Differential phosphorylation of Serine2 and Serine5 within this hepta-peptide repeat of CTD, affects transcription by coordinating the recruitment of histone methylation complexes and RNA processing factors. The Kin28 subunit of TFIIH catalyzes Ser-5 phosphorylation at initiation, whereas Ctk1 catalyzes Ser-2 phosphorylation during elongation. Recycling of RNAP II requires CTD dephosphorylation and Fcp1 has been identified as a CTD phosphatase with specificity for Ser-2 in vivo. Here we report that Ssu72, recently shown to be a component of the CPF complex of the pre-mRNA 3'-processing machinery, is a CTD phosphatase with specificity for Ser-5 in vitro and in vivo. Ssu72 catalyzes CTD S5-P dephosphorylation in association with the Pta1 component of the CPF complex, although its essential role in 3'-end processing is independent of catalytic activity. Depletion of Ssu72 impairs transcription in vitro and this defect can be rescued by recombinant, catalytically active Ssu72. We propose that Ssu72 has a dual role in transcription, one as a CTD S5-P phosphatase that regenerates the initiation-competent, hypophosphorylated form of RNAP II, and the other at the 3' end as a factor necessary for cleavage of pre-mRNA and efficient transcription termination.
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