Functional interactions between the transcription and mRNA 3' end processing machineries mediated by Ssu72 and Sub1.
Transcription and processing of pre-mRNA are coupled events. By using a combination of biochemical, molecular, and genetic methods, we have found that the phylogenetically conserved transcription factor Ssu72 is a component of the cleavage/polyadenylation factor (CPF) of Saccharomyces cerevisiae. Our results demonstrate that Ssu72 is required for 3' end cleavage of pre-mRNA but is dispensable for poly(A) addition and RNAP II termination. The in vitro cleavage defect caused by depletion of Ssu72 from cells can be rescued by addition of recombinant Ssu72. Ssu72 interacts physically and genetically with the Pta1 subunit of CPF. Overexpression of PTA1 causes synthetic lethality in an ssu72-3 mutant. Moreover, Sub1, which has been implicated in transcription initiation and termination, also interacts with Pta1, and overexpression of SUB1 suppresses the growth and processing defect of a pta1 mutation. Physical interactions of Ssu72 and Sub1 with Pta1 are mutually exclusive. Based on the interactions of Ssu72 and Sub1 with both the Pta1 of CPF and the TFIIB component of the initiation complex, we present a model describing how these novel connections between the transcription and 3' end processing machineries might facilitate transitions in the RNAP II transcription cycle.
He
Xiaoyuan
X
Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.
Khan
Asad U
AU
Cheng
Hailing
H
Pappas
Donald L
DL
Hampsey
Michael
M
Moore
Claire L
CL
D020413:3' Untranslated Regions / Q000235:genetics / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D002846:Chromatography, Affinity; D017931:DNA Primers / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics; D066298:In Vitro Techniques; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D010749:Phosphoprotein Phosphatases; D011061:Poly A; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D012319:RNA Polymerase II / Q000378:metabolism*; D012322:RNA Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D013489:Suppression, Genetic; D013728:Terminator Regions, Genetic; D015534:Trans-Activators / Q000378:metabolism; D035581:Transcription Factor TFIIB / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D039221:mRNA Cleavage and Polyadenylation Factors / Q000737:chemistry / Q000378:metabolism
D020413:3' Untranslated Regions; D002352:Carrier Proteins; D017931:DNA Primers; C076229:PTA1 protein, S cerevisiae; D012322:RNA Precursors; D011993:Recombinant Fusion Proteins; C416934:SSU72 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D035581:Transcription Factor TFIIB; D039221:mRNA Cleavage and Polyadenylation Factors; D011061:Poly A; D012319:RNA Polymerase II; D010749:Phosphoprotein Phosphatases