Anhidrotic ectodermal dysplasia (EDA) is a disorder characterized by poor development of hair, teeth, and sweat glands, and results from lesions in the X-linked EDA gene. We have cloned a 1.6-kilobase 5'-flanking region of the human EDA gene and used it to analyze features of transcriptional regulation. Primer extension analysis located a single transcription initiation site 264 base pairs (bp) upstream of the translation start site. When the intact cloned fragment or truncated derivatives were placed upstream of a reporter luciferase gene and transfected into a series of cultured cells, expression comparable with that conferred by an SV40 promoter-enhancer was observed. The region lacks a TATA box sequence, and basal transcription from the unique start site is dependent on two binding sites for the Sp1 transcription factor. One site lies 38 bp 5' to the transcription start site, in a 71-bp sequence that is sufficient to support up to 35% of maximal transcription. The functional importance of the Sp1 sites was demonstrated when cotransfection of an Sp1 expression vector transactivated the EDA promoter in the SL2 Drosophila cell line that otherwise lacks endogenous Sp1. Also, both Sp1 binding sites were active in footprinting and gel shift assays in the presence of either crude HeLa cell nuclear extract or purified Sp1 and lost activity when the binding sites were mutated. A second region involved in positive control was localized to a 40-bp sequence between -673 and -633 bp. This region activated an SV40 minimal promoter 4- to 5-fold in an orientation-independent manner and is thus inferred to contain an enhancer region.
"Both Wnt and Eda signaling are required for the formation of the first wave of hair and feather follicles and in tooth development, but the precise connection between these two signaling pathways, and other intercellular signaling events, is still unclear . In vitro experiments have suggested that Eda is directly regulated by Wnt signaling in the tooth bud (Pengue et al., 1999; Laurikkala et al., 2001; Durmowicz et al., 2002). "
[Show abstract][Hide abstract] ABSTRACT: To provide a genetic framework for investigating changes in airway submucosal gland function in human respiratory disease, we have investigated their counterparts in normal and mutant mice. We describe their morphogenesis in relation to the expression of genes encoding conserved intercellular signaling pathways. Submucosal glands are severely reduced in number and size in mice heterozygous for Fgf10. Glands are completely absent in mice lacking Ectodysplasin (Eda) and Edaradd (Eda receptor adaptor protein), members of the tumor necrosis (TNF) superfamily of signaling factors. Furthermore, components of the Eda and closely related pathways are transcribed throughout the respiratory system in the adult mouse. Finally, the temporal and spatial pattern of Bmp4 expression suggests that it may control submucosal gland development and homeostasis. Taken together, our observations have important implications for the better understanding of the submucosal gland remodeling that occurs in human respiratory disease.
"With the exception of the missing exon 2, the observed genomic organization of the ED1 gene in cattle was very similar to the human ED1 gene (Bayés et al. 1998) and the mouse ta gene (Srivastava et al. 1997). The transcription start site of the bovine ED1 gene was tentatively assigned by sequence alignment with the human ED1 sequence (Pengue et al. 1999). The region immediately surrounding the transcription start site is highly conserved between human and cattle. "
[Show abstract][Hide abstract] ABSTRACT: Anhidrotic ectodermal dysplasia (ED1) is characterized by hypotrichosis, reduced number of sweat glands, and incisior anodontia in human, mouse, and cattle. In affected humans and mice, mutations in the ED1 gene coding for ectodysplasin 1 are found. Ectodysplasin 1 is a novel trimeric transmembrane protein with an extracellular TNF-like signaling domain that is believed to be involved in the formation of hair follicles and tooth buds during fetal development. We report the construction of a 480-kb BAC contig harboring the complete bovine ED1 gene on BTA Xq22-Xq24. Physical mapping and sequence analysis of the coding parts of the ED1 gene revealed that a large genomic region including exon 3 of the ED1 gene is deleted in cattle with anhidrotic ectodermal dysplasia in a family of German Holstein cattle with three affected maternal half sibs.
Genome Research 11/2001; 11(10):1699-705. DOI:10.1101/gr.182501 · 14.63 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The RET proto-oncogene plays an important role in the initiation and progression of tumors derived from the neural crest. The cis-regulatory elements responsible for RET basal promoter activity have not been identified. To characterize these elements, a RET promoter DNA fragment (-453 to +227bp) was fused to a luciferase reporter and introduced into TT, a neural crest-derived cell line. Sequential 5' deletions of the promoter revealed that optimal expression of the RET promoter in TT cells required only 70bp of sequence upstream of the transcription start site, and contains two Sp1 binding sites. DNase I footprinting, electrophoretic mobility shift analysis (EMSA), and supershift assays revealed that this region binds both Sp1 and its related protein, Sp3. Additionally, RET basal promoter activity was abrogated by removal of these Sp1/Sp3 binding sites. The proximal two GC boxes were sufficient to allow transactivation of the RET promoter in Drosophila SL2 cells. Sp3 expression in these cells caused an additional activation of the promoter. These results demonstrate that the transactivation of the RET promoter within a neural crest-derived cell line is dependent on Sp1 and Sp3.
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