Hirschsprung disease (HSCR), or aganglionic megacolon, is a relatively common, multifactorial birth defect associated with the lack of intrinsic ganglion cells in the myenteric (Auerbach) and submucosal (Meissner) plexuses in the gastrointestinal tract. Clinically, the symptoms range from mild to severe and involve aganglionosis of the transverse colon and beyond (long segment HSCR), the splenic flexure and the descending colon (short segment HSCR), or the sigmoid colon only (classic HSCR). Research support by this grant has led to the mapping and identification of three genes that lead to HSCR susceptibility, namely, the genes for the receptor tyrosine kinase RET, the endothelin receptor B (ENDRB), and its physiological ligand endothelin-3 (EDN3). In the mouse, Ece1 (endothelin converting enzyme-1) has been identified as an additional HSCR gene, and the Dom mutation, also leading to aganglionosis, is close to being cloned. HSCR in a genetically heterogeneous multifactorial disorder, and additional susceptibility genes remain to be identified. The investigators' evidence indicates that HSCR is oligogenic requiring multiple, interacting disease susceptibility alleles for phenotypic expression. This hypothesis forms the basis of this proposal. To enable the genetic dissection of HSCR the investigators propose two major studies. First, they shall pursue the genetic mapping, identification, mutation and functional analyses of additional HSCR susceptibility genes, as well as the known HSCR genes, to distinguish between the hypotheses of non-allelic heterogeneity and multigenic inheritance. Second, genetic and biochemical methods will be used to assess the importance of genetic interactions between HSCR susceptibility genes in determining phenotypic expression. In particular, using the known HSCR genes, haplotype analysis in HSCR families, penetrance analysis in mouse models of aganglionosis segregating null or missense mutations, and in vitro yeast two-hybrid assays using normal and mutant proteins will be used to test known, and identify new, susceptibility genes for interactions. The long-term objective of this renewal proposal is to understand ganglion cell differentiation and development in the gut. More generally, the aim is to develop a paradigm for the genetic and molecular analysis of complex, multifactorial human diseases.
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