THE WINGED HELIX TRANSCRIPTION FACTOR HFH-4 IS EXPRESSED DURING CHOROID-PLEXUS EPITHELIAL DEVELOPMENT IN THE MOUSE EMBRYO

Mammalian hepatocyte nuclear factor-3 (HNF-3) and the Drosophila homeo tic gene fork head proteins are prototypes of an extensive family of c ell-specific transcription factors that share homology In the winged h elix DNA-binding domain. One of these mammalian family members, HNF-3/ fork head homolog-4 (HFH-4), was isolated by PCR amplification of rode nt brain cDNA and exhibits abundant expression in the adult bronchiola r epithelium. In this study, we performed in situ hybridization of sta ge-specific mouse embryos and report on a novel expression pattern of the HFH-4 gene in both the presumptive and differentiated choroid plex us epithelium, which is responsible for the synthesis and secretion of cerebrospinal fluid (CSF) proteins. We also showed that HFH-4 is a po tent transcriptional activator in cotransfection assays and defined se veral protein sequences important for HFH-4 transcriptional activity. We used in vitro DNA-binding site selection with recombinant HFH-4 pro tein and determined that the HFH-4 protein recognizes the DNA consensu s sequences HWDTGTTTGTTTA or KTTTGTTGTTKTW (where H is not G, W is A o r T, D is not C, and K is G or T). We used this HFH-4 consensus to ide ntify potential HFH-4 target gents in the choroid plexus epithelium an d demonstrated that these promoter sequences bind to recombinant HFH-4 protein in electrophoretic mobility shift assays, Recombinant HFH-4 f ormed specific protein-DNA completes with the promoter regions of the human prothrombin, beta amyloid precursor protein, alpha 1-antichymotr ypsin, cystic fibrosis transmembrane conductance regulator and rodent alpha 2-macroglubulin, growth hormone receptors, and insulin-like grow th factor II genes. Furthermore, me identified putative HFH-4 target g enes in the bronchiolar epithelium including the clara cell secretory protein gene and the HNF-3 alpha gene, a winged helix family member in volved in the transcriptional regulation of genes in the bronchiolar e pithelium. In support of these binding studies, cotransfection assays show that HFH-4 potentiates expression of the INF-3 alpha and clara ce ll secretory protein promoter regions.