title: The Not5 subunit of the ccr4-not complex connects transcription and translation.
abstract: Recent studies have suggested that a sub-complex of RNA polymerase II composed of Rpb4 and Rpb7 couples the nuclear and cytoplasmic stages of gene expression by associating with newly made mRNAs in the nucleus, and contributing to their translation and degradation in the cytoplasm. Here we show by yeast two hybrid and co-immunoprecipitation experiments, followed by ribosome fractionation and fluorescent microscopy, that a subunit of the Ccr4-Not complex, Not5, is essential in the nucleus for the cytoplasmic functions of Rpb4. Not5 interacts with Rpb4; it is required for the presence of Rpb4 in polysomes, for interaction of Rpb4 with the translation initiation factor eIF3 and for association of Rpb4 with mRNAs. We find that Rpb7 presence in the cytoplasm and polysomes is much less significant than that of Rpb4, and that it does not depend upon Not5. Hence Not5-dependence unlinks the cytoplasmic functions of Rpb4 and Rpb7. We additionally determine with RNA immunoprecipitation and native gel analysis that Not5 is needed in the cytoplasm for the co-translational assembly of RNA polymerase II. This stems from the importance of Not5 for the association of the R2TP Hsp90 co-chaperone with polysomes translating RPB1 mRNA to protect newly synthesized Rpb1 from aggregation. Hence taken together our results show that Not5 interconnects translation and transcription.
authors.affiliation: Department of Microbiology and Molecular Medicine, University of Geneva, Faculty of Medicine, Geneva, Switzerland; Institute of Genetics and Genomics of Geneva, Geneva, Switzerland.
authors.forename: Zoltan
authors.initials: Z
authors.lastname: Villanyi
authors.affiliation: Department of Microbiology and Molecular Medicine, University of Geneva, Faculty of Medicine, Geneva, Switzerland; Institute of Genetics and Genomics of Geneva, Geneva, Switzerland.
authors.forename: Virginie
authors.initials: V
authors.lastname: Ribaud
authors.affiliation: Department of Microbiology and Molecular Medicine, University of Geneva, Faculty of Medicine, Geneva, Switzerland; Institute of Genetics and Genomics of Geneva, Geneva, Switzerland.
authors.forename: Sari
authors.initials: S
authors.lastname: Kassem
authors.affiliation: Department of Microbiology and Molecular Medicine, University of Geneva, Faculty of Medicine, Geneva, Switzerland; Institute of Genetics and Genomics of Geneva, Geneva, Switzerland.
authors.forename: Olesya O
authors.initials: OO
authors.lastname: Panasenko
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Szeged, Szeged, Hungary.
authors.forename: Zoltan
authors.initials: Z
authors.lastname: Pahi
authors.affiliation: European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany.
authors.forename: Ishaan
authors.initials: I
authors.lastname: Gupta
authors.affiliation: European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany.
authors.forename: Lars
authors.initials: L
authors.lastname: Steinmetz
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Szeged, Szeged, Hungary.
authors.forename: Imre
authors.initials: I
authors.lastname: Boros
authors.affiliation: Department of Microbiology and Molecular Medicine, University of Geneva, Faculty of Medicine, Geneva, Switzerland; Institute of Genetics and Genomics of Geneva, Geneva, Switzerland.
authors.forename: Martine A
authors.initials: MA
authors.lastname: Collart
mesh_terms: D002467:Cell Nucleus / Q000235:genetics; D003593:Cytoplasm / Q000235:genetics; D039621:Eukaryotic Initiation Factor-3 / Q000235:genetics; D015966:Gene Expression Regulation, Fungal; D011132:Polyribosomes / Q000235:genetics; D014176:Protein Biosynthesis; D012319:RNA Polymerase II / Q000235:genetics; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics*; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D039621:Eukaryotic Initiation Factor-3; C111175:NOT5 protein, S cerevisiae; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D012319:RNA Polymerase II; C511736:RPB1 protein, S cerevisiae; C423309:RPB4 protein, S cerevisiae; C470553:RPB7 protein, S cerevisiae