MOLECULAR/ GENETIC BASIS OF EARLY CONOTRUNCAL MORPHOGENESIS

22q11.2的微缺失与超过90%的DiGeorge综合征病例相关，被称为DiGeorge关键区（DGCR），其大小变化很大，不能预测表型的严重程度。DGCR中有一个200- 250kb的片段，是所有缺失患者共有的，被称为最小迪乔治临界区（MDGCR），这表明至少有一些与DGS表型相关的基因存在于该区域。为了验证这一假设，研究人员组装了小鼠16号染色体的cosmid和BAC组，并建立了一个与MDGCR同步的区域（Galili et al., 1997）。这已经扩展到包括与大多数2.5 Mb DGCR合成的区域，并包含人类DGCR中已知的25或26个基因。小鼠和人DGCR在基因含量、序列和内含子/外显子组成方面几乎完全一致，包括6个对流出道形态发生可能重要的基因。确定这些基因和其他基因在流出道形态发生中的作用的方法是基于以下假设：(1)迪乔治表型是DGCR内一个或多个基因拷贝数变化的结果；(2)关键的形态发生事件可能依赖于一个以上基因的协调表达；(3) DGS表型的不同程度是不同基因组合表达变化以及遗传背景变化的结果；(4)在MDGCR中至少有一个基因对早期心脏形态发生的某些方面至关重要。因此，我们的目标是利用过去四年中产生的试剂和序列数据来评估单个基因的形态发生功能，利用转基因动物重叠BACs和cosmids来评估多个基因在DGCR内“原生”遗传环境下的形态发生作用。并探索研究人员通过代表性差异分析和酵母2杂交系统在早期心脏形态发生和随后在“非缺失”患者先天性流出道异常中的作用所确定的新基因的作用。

(Adapted from the Applicant's Abstract) The microdeletion of 22q11.2 associated with over 90 percent of the cases of DiGeorge syndrome, referred to as the DiGeorge Critical Region (DGCR), is highly variable in size and does not predict severity of phenotype. Within the DGCR is a 200-250 kb segment common to all deleted patients designated the minimal DiGeorge Critical Region (MDGCR), suggesting that at least some genes relevant to the DGS phenotype reside in this region. To test this hypothesis, the investigators assembled a cosmid and BAC contig of mouse chromosome 16 and established a region of synteny with the MDGCR (Galili et al., 1997). This has been extended to include regions syntenic to most of the 2.5 Mb DGCR, and contains 25 or the 26 known genes within the human DGCR. Synteny between mouse and human DGCR is almost complete with respect to gene content, order and intron/exon composition, and includes six genes of potential importance to outflow tract morphogenesis. The approach to determining the role of these and other genes to outflow tract morphogenesis is based upon the hypothesis that: (1) the DiGeorge phenotype is the result of a change in copy number of one or more genes within the DGCR; (2) the critical morphogenetic event(s) may depend upon the coordinated expression of more than one gene; (3) the variable severity of DGS phenotypes is the result of changes in expression of different combinations of genes as well as variable genetic backgrounds; (4) within the MDGCR is at least one gene critical to some aspect of early heart morphogenesis. The goals are, therefore to evaluate the morphogenetic function of individual genes using reagents and sequence data generated over the past four years, use animals transgenic for overlapping BACs and cosmids to evaluate the morphogenetic role of multiple genes in the context of their "native" genetic environment within the DGCR, and explore the role of new genes the investigators have identified by representational difference analysis and with the yeast 2-hybrid system in early heart morphogenesis and subsequent role in congenital outflow tract abnormalities within "non-deleted" patients.