title: CCAAT binding NF-Y-TBP interactions: NF-YB and NF-YC require short domains adjacent to their histone fold motifs for association with TBP basic residues.
abstract: Both the TATA and CCAAT boxes are widespread promoter elements and their binding proteins, TBP and NF-Y, are extremely conserved in evolution. NF-Y is composed of three subunits, NF-YA, NF-YB and NF-YC, all necessary for DNA binding. NF-YB and NF-YC contain a putative histone-like motif, a domain also present in TBP-associated factors (TAFIIs) and in the subunits of the transcriptional repressor NC2. Immunopurification of holo-TFIID with anti-TBP and anti-TAFII100 antibodies indicates that a fraction of NF-YB associates with TFIID in the absence of NF-YA. Sedimentation velocity centrifugation experiments confirm that two pools of NF-YB, and most likely NF-YC, exist: one associated with NF-YA and binding to the CCAAT box; another involved in high molecular weight complexes. We started to dissect NF-Y-TFIID interactions by showing that: (i) NF-YB and NF-YC interact with TBP in solution, both separately and once bound to each other; (ii) short stretches of both NF-YB and NF-YC located within the evolutionary conserved domains, adjacent to the putative histone fold motifs, are necessary for TBP binding; (iii) TBP single amino acid mutants in the HS2 helix, previously shown to be defective in NC2 binding, are also unable to bind NF-YB and NF-YC.
authors.affiliation: Dipartimento di Genetica e Biologia dei Microrganismi, Università di Milano, Via Celoria 26, 20133 Milano, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Bellorini
authors.forename: D K
authors.initials: DK
authors.lastname: Lee
authors.forename: J C
authors.initials: JC
authors.lastname: Dantonel
authors.forename: K
authors.initials: K
authors.lastname: Zemzoumi
authors.forename: R G
authors.initials: RG
authors.lastname: Roeder
authors.forename: L
authors.initials: L
authors.lastname: Tora
authors.forename: R
authors.initials: R
authors.lastname: Mantovani
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D023081:CCAAT-Binding Factor / Q000378:metabolism*; D022762:CCAAT-Enhancer-Binding Proteins; D002499:Centrifugation, Density Gradient; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli; D006657:Histones; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017510:Protein Folding; D051381:Rats; D016385:TATA Box; D035181:TATA-Box Binding Protein; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000378:metabolism*; D019473:Transcription Factors, TFII / Q000378:metabolism
chemical_list: D023081:CCAAT-Binding Factor; D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D006657:Histones; C000593008:Nfyc protein, rat; D035181:TATA-Box Binding Protein; D035362:Transcription Factor TFIID; D014157:Transcription Factors; D019473:Transcription Factors, TFII