Dissection of the NF-Y transcriptional activation potential.
NF-Y is a trimeric CCAAT-binding factor with histone fold subunits (NF-YB/NF-YC) and bipartite activation domains located on NF-YA and NF-YC. We reconstituted the NF-Y activation potential in vivo with GAL4 DBD fusions. In the GAL4-YA configuration, activation requires co-expression of the three subunits; with GAL4-YB and GAL4-YC, transfections of the histone fold partners are sufficient, provided that the Q-rich domain of NF-YC is present. Combinations of mutants indicate that the Q-rich domains of NF-YA and NF-YC are redundant in the trimeric complex. Glutamines 101 and 102 of NF-YA are required for activity. We assayed NF-Y on different promoter targets, containing single or multiple GAL4 sites: whereas on a single site NF-Y is nearly as powerful as VP16, on multiple sites neither synergistic nor additive effects are observed. NF-Y activates TATA and Inr core elements and the overall potency is in the same range as other Q-rich and Pro-rich activation domains. These results represent the first in vivo evidence of subunit interactions studies and further support the hypothesis that NF-Y is a general promoter organizer rather than a brute activator.
de Silvio
A
A
Dipartimento di Genetica e Biologia dei Microrganismi, Universitá di Milano, Via Celoria 26, 20133 Milano, Italy.
Imbriano
C
C
Mantovani
R
R
D016475:3T3 Cells; D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics; D051379:Mice; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors