NF-Y binds to the CCAAT box motif of the FGF-4 gene and promotes FGF-4 expression in embryonal carcinoma cells

Abstract

FGF‐4 appears to be the first fibroblast growth factor (FGF) expressed during embryogenesis, and its expression is critical for early mammalian development. FGF‐4 is expressed in the embryonic cell lines, F9, D3, and NT2/D1; but its expression in these cells is repressed upon differentiation. Transcription of the FGF‐4 gene in embryonic cells is regulated by an enhancer in the third exon and by a positive regulatory region upstream of the transcription start site. A CCAAT box motif within the positive regulatory region has been shown to support FGF‐4 expression, but the factor that binds to this site in vivo has not been identified. In this report, we demonstrate that the transcription factor complex NF‐Y binds to the FGF‐4 CCAAT box motif when nuclear extracts from each of the embryonic cell lines and their differentiated cells were examined by gel mobility shift analyses. Importantly, we demonstrate that expression of a dominant‐negative NF‐YA mutant protein reduces the expression of FGF‐4 promoter/reporter gene constructs in F9 EC cells. Hence, we provide strong evidence that the transcription factor NF‐Y is involved in the expression of the FGF‐4 gene. Mol. Reprod. Dev. 48:301–309, 1997. © 1997 Wiley‐Liss, Inc.

The CCAAT box binding transcription factor, nuclear factor-Y (NF-Y) regulates transcription of human aldo-keto reductase 1C1 (AKR1C1) gene

“…6B). Various other authors have also observed a decrease in promoter activity of the target gene upon co-expression with the dominant-negative NF-YA mutant (Lamb et al, 1997; Liu et al, 1997; Sugiura and Takishima, 2000; Kam et al, 2005; Wageningen et al, 2008). …”

Isolation, characterization, and differential expression of the murine Sox-2 promoter

“…Interestingly, the FGF-4, TβR-II, and CYP1A1 genes each contain identical CCAAT box-like sequence motifs in each of their respective promoters (5′-TGATTGGCAG-3′). Unlike the Sox-2 CCAAT box motif, gel mobility shift analysis of the FGF-4, TβR-II, and CYP1A1 CCAAT box motifs revealed formation of two distinct DNA/protein complexes, one of which contains NF-Y (Boucher et al, 1993(Boucher et al, , 1995Lamb et al, 1997;Kelly et al, 1998). Both complexes are specific for the CCAAT box-like motif, as an unlabeled FGF-4, TβR-II, or CYP1A1 mutant CCAAT box competitor, in which only the CCAAT box has been scrambled, does not compete for either complex (Kelly and Rizzino, unpublished data).…”

“…These genes all contain the sequence (5′-TGATTGGC AG-3′), which differs by only one base pair (underlined) with the Sox-2 CCAAT box (5′-TGATTGGCCG-3′). Previous gel mobility shift assays demonstrated that the CCAAT box motif from each of these three genes can form two DNA/protein complexes, one of which contains NF-Y (Boucher et al, 1993(Boucher et al, , 1995Lamb et al, 1997;Kelly et al, 1998). To determine whether NF-Y binds the Sox-2 CCAAT box motif, we performed gel mobility shift analysis using a dsODN probe containing the CCAAT box sequence of the Sox-2 promoter.…”

Transcriptional Activation of the Type II Transforming Growth Factor-β Receptor Gene upon Differentiation of Embryonal Carcinoma Cells

“…NF-Y was originally identified as a ubiquitously expressed protein that binds to the Y box motif, originally defined as an inverted CCAAT box motif (CTGATTGGYY) in all MHC class II genes that is critical for tissue specific gene expression (71). Although the exact mechanism(s) by which NF-Y regulates transcription is unclear, it has been demonstrated to act as both a positive (66,72,73) and a negative (74) transcription factor. Several studies suggest that NF-Y acts by stabilizing or recruiting the binding of additional factors to adjacent promoter elements (75,76).…”