title: Repression of promoter activity by CNOT2, a subunit of the transcription regulatory Ccr4-not complex.
abstract: The evolutionary conserved Ccr4-Not complex controls mRNA metabolism at multiple levels in eukaryotic cells. Genetic analysis of not mutants in yeast identifies a negative role in transcription, which is dependent on core promoter structure. To obtain direct support for this we targeted individual core subunits of the human Ccr4-Not complex to promoters in transient transfections of human cells. In this experimental setup we found that the CNOT2 and CNOT9(hRcd1/hCaf40) subunits act as repressors of reporter gene activity. Interestingly, recruitment of other Ccr4-Not subunits did not affect the reporter gene. The major repression function of CNOT2 is localized in a specialized protein motif, the Not-Box. This conserved motif is present in all CNOT2 orthologs and surprisingly also in CNOT3 orthologs. Repression by the Not-Box was sensitive to treatment with the histone deacetylase inhibitor trichostatin A. In addition, mutation of a canonical TATA-box enhanced repression. Our experiments show for the first time direct regulation of promoter activity by components of the Ccr4-Not complex.
authors.affiliation: Department of Physiological Chemistry, Stratenum STR 3.229, University Medical Center Utrecht, Universiteitsweg 100, Utrecht 3584 CG, The Netherlands.
authors.forename: Carin G M
authors.initials: CG
authors.lastname: Zwartjes
authors.forename: Sandrine
authors.initials: S
authors.lastname: Jayne
authors.forename: Debbie L C
authors.initials: DL
authors.lastname: van den Berg
authors.forename: H T Marc
authors.initials: HT
authors.lastname: Timmers
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology*; D017930:Genes, Reporter; D006801:Humans; D008969:Molecular Sequence Data; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000737:chemistry / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000737:chemistry / Q000502:physiology*
chemical_list: C485365:CNOT2 protein, human; D004268:DNA-Binding Proteins; C539882:NR4A2 protein, human; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D012097:Repressor Proteins; D014157:Transcription Factors