Molecular Genetics of X-linked Cataracts

先天性白内障，在临床上被定义为出生时就存在的白内障，是由 正常的晶状体发育所需的基因。这项建议的主要目标是确定 先天性白内障--人类X-连锁白内障牙科综合征的基因缺陷 与小角膜和牙齿畸形有关。根据X染色体上的表型和位置， XCAT小鼠是人类XLCD最好的动物模型。我们已经通过一个 种间回交，鉴定了两个标记DxWas31和DxPas18，它们没有与 XCAT。筛选酵母人工染色体(YAC)和细菌人工染色体(BAG)文库 利用这些标记构建了XCAT关键区的900kb重叠群图。XCAT关键区域将是 通过袋子转基因互补实验精炼，旨在识别含有以下物质的单个袋子 XCAT基因。我们将获得这个袋子的序列，并分析它的潜在转录单位和EX 候选的xCAT基因将通过南方动物园的印迹和数据库来筛选其在晶状体中的表达 寻找跨物种的保护。在产前保守和表达的候选基因 将评估晶状体在xCAT小鼠中的表达。他们还将接受XCAT基因突变的评估。 突变的xCAT基因与白内障表型的相关性将通过靶向研究来证实。 在转基因动物中中断xCAT。最后，将使用正常的小鼠xCAT序列来克隆 相应的正常人类基因。在确定了正常XLCD基因的结构后， XLCD患者将接受检查，以确定是否存在xCAT人类同源基因突变。识别 XLCD基因将有助于我们理解晶状体中涉及的分子事件 发展。

Congenital cataracts, clinically defined as the presence of cataracts at birth, are the result of mutations in genes that are required for,proper lens development. The primary objective of this proposal is to identify the gene defective in the human X-linked cataract dental (XLCD) syndrome, a type of congenital cataract associated with microcornea and dental anomalies. Based on phenotype and location on the X chromosome, the Xcat mouse is the best animal model for human XLCD. We have positionally mapped Xcat through an interspecific backcross and identified two markers DxWas31 and DxPas18 that show no recombination with Xcat. Yeast artificial chromosome (YAC) and bacterial artificial chromosome (BAG) libraries were screened with these markers to develop a 900kb contig map of the Xcat critical region. The Xcat critical region will be refined through BAG transgenic complementation experiments designed to identify a single BAG that contains the Xcat gene. We will obtain the sequence of this BAG and analyze it for potential transcription units and ex Candidate Xcat genes will be screened for expression in the lens and by Southern zoo blots and database searches for conservation across species. Candidate genes that are conserved and expressed in prenatal lens will be evaluated for expression in Xcat mice. They will also be evaluated for mutations in the Xcat cDNA. The correlation between the mutant Xcat gene and the cataract phenotype will be confirmed by targeted disruption of Xcat in transgenic animals. Finally, the normal mouse Xcat sequence will be used to clone the corresponding normal human gene. After determining the structure of the normal XLCD gene, XLCD patients will be examined to determine if mutations in the Xcat human homolog are present. Identifying the XLCD gene would contribute to the our understanding of the molecular events involved in lens development.