Evolutionary Conserved Interaction between CstF-64 and PC4 Links Transcription, Polyadenylation, and Termination

Department of Biological Sciences, Columbia University, New York, NY 10027, USA.
Molecular Cell (Impact Factor: 14.02). 06/2001; 7(5):1013-23. DOI: 10.1016/S1097-2765(01)00236-2
Source: PubMed


Tight connections exist between transcription and subsequent processing of mRNA precursors, and interactions between the transcription and polyadenylation machineries seem especially extensive. Using a yeast two-hybrid screen to identify factors that interact with the polyadenylation factor CstF-64, we uncovered an interaction with the transcriptional coactivator PC4. Both human proteins have yeast homologs, Rna15p and Sub1p, respectively, and we show that these two proteins also interact. Given evidence that certain polyadenylation factors, including Rna15p, are necessary for termination in yeast, we show that deletion or overexpression of SUB1 suppresses or enhances, respectively, both growth and termination defects detected in an rna15 mutant strain. Our findings provide an additional, unexpected connection between transcription and polyadenylation and suggest that PC4/Sub1p, via its interaction with CstF-64/Rna15p, possesses an evolutionarily conserved antitermination activity.

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    • "Moreover, according to our protein–protein interaction analysis, TvCstF77, and TvCstF64 interact with TvCPSF160 and TvPC4, respectively ; for this reason these proteins were included in the model. It important to mention that these interactions were previously reported (Calvo and Manley 2001; Zhao et al. 1999). Since Clp1 interacts with PAP and CFI25 (de Vries et al. 2000), so TvClp1 was included in the model. "
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    ABSTRACT: Trichomonas vaginalis is the causative agent of trichomonosis, a sexually transmitted disease (STD) that affects over 180 million people worldwide. This parasite is capable to infect the urogenital tract of women and men, both microenvironments might affect the expression of key genes that may be involved in the parasite pathogenesis. The processing of 3′ end of mRNA promotes mRNA stability in many eukaryotes, however in T. vaginalis this molecular machinery is under research. By means of an in silico analysis we identified putative proteins of the 3′ end mRNA processing machinery of T. vaginalis, and by RT-PCR assays we evaluated the expression of eight of these genes in a female and male T. vaginalis isolates. According to the in silico analysis, the T. vaginalis 3′ end mRNA processing machinery, comprises a similar complex and protein factors that those described in Homo sapiens, Arabidopsis thaliana, Saccharomyces cerevisiae and Entamoeba histolytica. The complex contains several sub-complexes, including cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), cleavage factor I (CFIm) and cleavage factor II (CFIIm). We demonstrated that genes tvpsf2p, tvcfi25, tvcpsf160, tvcpsf73, tvfip1, tvpap1, tvpc4 and tvpabp are expressed in male or female T. vaginalis isolates. Besides we identify two different isoforms of TvPC4. T. vaginalis genome contains most of genes encoding for 3′ end mRNA processing, which may be transcriptionally active and could be involved in the capping, splicing, cleavage and polyadenylation of mRNAs in this parasite. Further studies are necessary to elucidate the biological meaning of our findings.
    Full-text · Article · Feb 2015 · Genes & genomics
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    • "Twenty-five milliliters of culture at an OD 600 of 0.3–4 growing at 30ºC and induced with 2% galactose was permeabilized and nascent RNA labeled with [α- 32 P]UTP for 5 min at 30ºC as described (Birse et al., 1998; Calvo and Manley, 2001). After partial hydrolysis, RNA was hybridized directly to filter immobilized YLR454 probes. "
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    ABSTRACT: The transcriptional coactivator Sub1 has been implicated in several steps of mRNA metabolism in yeast, such as the activation of transcription, termination and 3'-end formation. In addition, Sub1 globally regulates RNA polymerase II phosphorylation, and most recently it has been shown that it is a functional component of the PIC. Here, we present evidence that Sub1 plays a significant role in transcription elongation by RNA polymerase II. We show that SUB1 genetically interacts with the gene encoding the elongation factor Spt5, that Sub1 influences Spt5 phosphorylation by the CTD kinase Bur1, and that both Sub1 and Spt5 copurify in the same complex, likely during early transcription elongation. Indeed, our data indicate that Sub1 influences Spt5-Rpb1 interaction. In addition, biochemical and molecular data show that Sub1 influences transcription elongation of constitutive and inducible genes, and that it associates with coding regions in a transcription-dependent manner. Altogether our results indicate that Sub1 associates with Spt5, influences Spt5-Rpb1 complex levels and consequently transcription elongation rate.
    Preview · Article · Sep 2012 · Molecular biology of the cell
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    • "The allosteric model proposes that the presence of a polyadenylation sequence on the RNA triggers a change in the factors associated with the polymerase [26]. In this model, binding of the cleavage and polyadenylation specificity factors (CPSFs) and the cleavage stimulation factors (CstFs) to the AAUAAA polyadenylation signal on the nascent pre-mRNA favors transcription termination by displacing elongation factors and consequently rendering Pol II less processive [24], [27], [28], [29]. In the torpedo model, the cleavage event at the polyadenylation site generates a new 5′ end [30]. "
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