A multisubunit 3' end processing factor from yeast containing poly(A) polymerase and homologues of the subunits of mammalian cleavage and polyadenylation specificity factor

Department of Cell Biology, Biozentrum, University of Basel, Switzerland.
The EMBO Journal (Impact Factor: 10.75). 09/1997; 16(15):4727-37. DOI: 10.1093/emboj/16.15.4727
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ABSTRACT Polyadenylation is the second step in 3' end formation of most eukaryotic mRNAs. In Saccharomyces cerevisiae, this step requires three trans-acting factors: poly(A) polymerase (Pap1p), cleavage factor I (CF I) and polyadenylation factor I (PF I). Here, we describe the purification and subunit composition of a multiprotein complex containing Pap1p and PF I activities. PF I-Pap1p was purified to homogeneity by complementation of extracts mutant in the Fip1p subunit of PF I. In addition to Fip1p and Pap1p, the factor comprises homologues of all four subunits of mammalian cleavage and polyadenylation specificity factor (CPSF), as well as Ptalp, which previously has been implicated in pre-tRNA processing, and several as yet uncharacterized proteins. As expected for a PF I subunit, pta1-1 mutant extracts are deficient for polyadenylation in vitro. PF I also appears to be functionally related to CPSF, as it polyadenylates a substrate RNA more efficiently than Pap1p alone. Possibly, the observed interaction of the complex with RNA tethers Pap1p to its substrate.

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Available from: Walter Keller, Aug 19, 2015
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    • "The plant ortholog of WDR33, the protein FY, has been genetically shown to be involved in 39 processing (Simpson et al. 2003) and is associated with other CPSF subunits (Herr et al. 2006; Hunt et al. 2008; Manzano et al. 2009). In S. cerevisiae, the orthologs of CPSF160, CPSF100, CPSF73, CPSF30, WDR33, and hFip1, together with poly(A) polymerase and Mpe1p, form the polyadenylation factor I (PF I) (Preker et al. 1997), which is part of a larger assembly (holo-CPF) (Nedea et al. 2003). The symplekin ortholog Pta1p is not part of PF I but mediates its association with the rest of holo-CPF. "
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    • "CPF is thought to define the cleavage site by recognizing short U-rich tracts that flank the poly(A) site. The upstream tract is generally 10 nt or closer to the cleavage site (Preker et al. 1997; Graber et al. 1999). Structural analysis of the ternary complex of Pap1, RNA, and ATP has shown that 4 nt are needed to reach from the active site of Pap1 to the surface of the enzyme (Balbo and Bohm 2007). "
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    • "Here, using genetic modifier screens in C. elegans, we have uncovered a role of the novel protein SYDN-1 and the NpolyA processing factor PFS-2 in neuronal development. The PFS2 (Polyadenylation factor subunit 2) protein family is characterized by the presence of seven highly conserved WD repeats (Smith et al., 1999), and was originally identified from yeast and mammalian cells based on the polyadenylation activity (Preker et al., 1997; Shi et al., 2009). Yeast Pfs2p directly interacts with proteins in subcomplexes of NpolyA and possibly acts as a scaffold for the 3Ј- end processing machinery (Ohnacker et al., 2000). "
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