X-linked anhidrotic ectodermal dysplasia (EDA) is the most frequently occurring of more than 175 ectodermal dysplasias affecting one or more skin appendages. The gene that is mutated to cause this disorder encodes a protein, which we have named ectodysplasin-A, has a single transmembrane region with collagenous and TNF-ligand segments in a long extracelliular carboxyterminal tail. Because individuals with EDA have sparse hair, rudimentary teeth, and few sweat glands, the gene is likely involved at an early point in development. We had earlier shown that the promoter region has transcription elements that include enhancers, possibly implicated in the tissue specificity of the gene; they include sites that implicate the Wnt and EGF pathways. We had also demonstrated that the Tabby mouse, which has many of the features observed in human EDA, is specifically mutated in the orthologous mouse gene. The gene was known to have 3 different classes of protein products (isoforms); we have now found additional isoforms for the functional A form. We have been investigating whether the different protein products function to help form different skin appendages, or whether they cooperate for all skin appendage formation. To distinguish among these alternatives, we put one of the isoforms of the gene back into Tabby animals as a transgene. The result is unequivocal: various skin appendages are restored at least partially by that isoform. We have now found that the same isoform has two kinds of effects in embryonic mice: determining the formation of certain appendages, including some types of hair follicles, and increasing the size of others, including sebaceous glands. We also have characterized eye phenotypes of Tabby mice including blindness and inflammation susceptibility, and they are also reversed by supplementation with the same isoform. We are now beginning to look at Tabby animals transgenic for other (isoforms), or for combinations of isoforms, turning their formation on or off at defined times during fetal development in the mice by using a controllable promoter. Several groups have shown that the first known EDA isoforms can bind to different cellular receptors to effect the control of gene expression by a transcription control system (NF-kB). We are now doing tests to identify the receptors for newly found EDA isoforms to determine the relative contribution of different isoforms to the formation and maintenance of the appendages, and the pathways involved. To extend the analyses, we have initiated extensive microarray gene expression profiling for skin appendages at various times during their development. The studies thus far implicate a small number of genes that show significant quantitative changes in expression in Tabby compared to wild-type mouse skin. They include several major signal transduction pathways such as Wnt, SHH, and lymphotoxin in conjunction with NF-kB; and the changes are largely restored to normal levels in animals in which a transgene has restored much of the skin appendages. In ongoing complementary studies, extensions of promoter analyses are designed to understand the selective maintenance of ectodysplasin at a critical level in target cells; and the combination of histology, phenotype analysis, and gene expression assays in transgenic animals are providing information about the function of different isoforms.
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