ANALYSIS OF DOMINANT MEGACOLON--ANOTHER MODEL FOR HIRSCHSPRUNG DISEASE

显性巨结肠的杂合子动物 (DOM/+)在神经脊发育中表现出多种缺陷 包括皮肤中黑素细胞数量的减少和 结肠内无肌间神经节。一种人类先天 先天性巨结肠症也表现为直肠绞痛 无神经节细胞增多症，并可与色素减退有关。因此， DOM/+小鼠，以及花斑和致命的斑点突变体， 作为这种疾病的小鼠模型。DOM出现了，并一直 维护在a C57BL/6JLe X C3HeB/FeJLe-a/a(B6C3F1)上 杂交背景，然而表型的严重性是 受遗传背景的影响很大。使用连续 对不同近交系小鼠的回交，我们发现 在DOM/C3系中，斑点和存活率都增加了 与DOM/B6行进行比较。使用亚种间和亚种内 交叉DOM间隔缩小到&lt；0.001厘米(2 重组子/1700个减数分裂)。一张物理地图已经组装好了 来自该地区，DOM被限制在~150 kb 细分市场。我们已经使用单遍排序来确定候选人 基因。对候选基因的分析导致了 分子缺陷的鉴定。评估…的表型 DOM/DOM纯合子小鼠，联系最紧密的标记是 适用于基因型杂交后代。我们发现， DOM基因抑制两个神经脊标记Trp2的表达 EDNRB将DOM基因放置在神经脊的早期 发展路径。未来在这一领域的研究将包括现场 用杂交分析来描述神经脊线的发育 DOM/DOM突变胚胎。我们的神经脊体外培养系统 并将使用聚合嵌合体分析来确定 DOM缺陷是神经脊来源的黑素细胞所固有的。 先天性巨结肠症与DOM关系的研究 将与Aravinda Chachravarti合作进行探索 (壁外)通过使用人类谱系分析。

Animals heterozygous for Dominant megacolon (Dom/+) exhibit multiple defects in neural crest development including reduced numbers of melanocytes in the skin and an absence of myenteric ganglion in the colon. A human congenital disorder, Hirschsprung disease also exhibits rectocolic aganglionosis and can be associated with hypopigmentation. Thus Dom/+ mice, as well as the piebald and lethal spotting mutants, serve as mouse models for this disease. Dom arose and has been maintained on a C57BL/6JLe X C3HeB/FeJLe-a/a (B6C3F1) hybrid background, however the severity of the phenotype is dramatically influenced by genetic background. Using continuous backcrossing to different inbred strains of mice, we have found that both spotting and survival are increased in Dom/C3 lines in comparison to Dom/B6 lines. Using inter- and intra-subspecific crosses the Dom interval was narrowed to < 0.001 cM (2 recombinants/1700 meioses). A physcial map has been assembled from the region and Dom has been restricted to an ~150 kb segment. We have used single pass sequencing to identify candidate genes. Analysis of the candidate genes has resulted in the identification of the molecular defect. To evaluate the phenotype of Dom/Dom homozygous mice, the most closely linked markers were applied to genotype intercross progeny. We have found that the DOM gene disrutps expression of two neural crest markers, TRP2 and EDNRB placing the Dom gene early in the neural crest development pathway. Future studies in this area will include in situ hybridization analyses to describe neural crest development in Dom/Dom mutant embryos. Our in vitro neural crest culture system and aggregation chimera analyses will be used to determine if the Dom defect acts intrinsic to the neural crest derived melanocytes. Investigation of the involvement of Dom in Hirschsprung disease will be explored in collaboration with Aravinda Chachravarti (Extramural) throught the use of human pedigree analysis.