title: NF-YC complexity is generated by dual promoters and alternative splicing.
abstract: The CCAAT box is a DNA element present in the majority of human promoters, bound by the trimeric NF-Y, composed of NF-YA, NF-YB, and NF-YC subunits. We describe and characterize novel isoforms of one of the two histone-like subunits, NF-YC. The locus generates a minimum of four splicing products, mainly located within the Q-rich activation domain. The abundance of each isoform is cell-dependent; only one major NF-YC isoform is present in a given cell type. The 37- and 50-kDa isoforms are mutually exclusive, and preferential pairings with NF-YA isoforms possess different transcriptional activities, with specific combinations being more active on selected promoters. The transcriptional regulation of the NF-YC locus is also complex, and mRNAs arise from the two promoters P1 and P2. Transient transfections, chromatin immunoprecipitations, and reverse transcription-PCRs indicate that P1 has a robust housekeeping activity; P2 possesses a lower basal activity, but it is induced in response to DNA damage in a p53-dependent way. Alternative promoter usage directly affects NF-YC splicing, with the 50-kDa transcript being excluded from P2. Specific functional inactivation of the 37-kDa isoform affects the basal levels of G(1)/S blocking and pro-apoptotic genes but not G(2)/M promoters. In summary, our data highlight an unexpected degree of complexity and regulation of the NF-YC gene, demonstrating the existence of a discrete cohort of NF-Y trimer subtypes resulting from the functional diversification of Q-rich transactivating subunits and a specific role of the 37-kDa isoform in suppression of the DNA damage-response under growing conditions.
authors.affiliation: Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano, Via Celoria 26, 20133 Milano, Italy.
authors.forename: Michele
authors.initials: M
authors.lastname: Ceribelli
authors.forename: Paolo
authors.initials: P
authors.lastname: Benatti
authors.forename: Carol
authors.initials: C
authors.lastname: Imbriano
authors.forename: Roberto
authors.initials: R
authors.lastname: Mantovani
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001483:Base Sequence; D023081:CCAAT-Binding Factor / Q000235:genetics* / Q000502:physiology*; D045744:Cell Line, Tumor; D019295:Computational Biology / Q000379:methods; D004249:DNA Damage; D019281:Dimerization; D006367:HeLa Cells; D006657:Histones / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
chemical_list: D023081:CCAAT-Binding Factor; D006657:Histones; C469358:NFYC protein, human; D020033:Protein Isoforms
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