Identifying Cardiomyopathy Genes in Mice and Drosophila

描述(由申请人提供)：遗传学方法，尤其是在既能准确测量心脏功能又能进行有效的基因筛查的模型系统中，对于在识别心力衰竭的遗传学基础方面取得实质性进展是必要的。在提案中，我们将检验这样一个假设，即强大的诱变剂N-乙基-N-亚硝基脲(ENU)，在小鼠中的诱变，以及在果蝇中的基因缺失筛选，将导致发现人类心力衰竭的新的致病和疾病修饰基因。为了验证这些假设，我们在8周龄和16周龄的成年小鼠中使用无创超声心动图进行了隐性突变筛查，以筛查心脏功能异常，并已经确定了1号染色体上映射到心肌病表型的可遗传区域。为了补充小鼠的研究，我们建议使用苍蝇来识别新的心肌病基因。果蝇遗传学提供了19,000多个插入整个果蝇基因组的分子定义的P-元件，促进了高密度基因组覆盖的产生。我们开发了一种创新的方法来研究成年清醒果蝇的表型心脏功能，并发现了导致扩张型心肌病的短原肠形成(SOG)基因中的一个P元素突变。基于我们的初步发现，我们认为小鼠和果蝇的遗传学可以识别导致人类扩张型心肌病的新基因和机制。因此，我们提出了以下具体目标：目标1：利用ENU表型驱动的隐性筛选，在成年小鼠中寻找导致心肌病的新基因。我们建议定位位于1号染色体上的心功能异常的ENU家族的致病基因。目的：探讨DPP/BMP信号通路突变导致成年果蝇扩张型心肌病的生化和遗传学机制。将在果蝇身上进行遗传互补实验，以证明SOG缺乏会导致心肌病。目的3：检测在压力超负荷条件下，小鼠BMP途径的拮抗性降低是否会导致心肌病的表型。TAG实验将在缺乏内源性哺乳动物BMP拮抗剂Chordin和Noggin的基因敲除小鼠身上进行。目的4：通过对果蝇进行全基因组缺失突变筛选，寻找导致心肌病的新基因。来自Exelixis收集的突变缺失突变的果蝇将使用光学相干断层扫描进行心肌病筛查，然后对致病基因进行精细定位(S)。因此，这四个综合目标将利用小鼠和果蝇遗传学的力量来识别和评估新的候选基因在心肌病中的作用。

DESCRIPTION (provided by applicant): Genetic approaches, preferentially in model systems that both allow accurate measurement of heart function as well as efficient genetic screening, are necessary for substantial progress in identifying the genetic basis of heart failure. In proposal, we will test the hypothesis that the powerful mutagen, N-ethyl-N- nitrosourea (ENU), mutagenesis in mice, and gene-deletion screens in Drosophila, will lead to the discovery of novel disease-causing and disease-modifying genes for human heart failure. In order to test these hypotheses, we have performed a recessive mutagenesis screen in adult mice at 8 and 16 weeks of age using non-invasive echocardiography to screen for abnormalities in cardiac function and have already identified a heritable region on chromosome 1 that maps to a cardiomyopathic phenotype. To complement the mouse studies we propose to use the fly to identify novel cardiomyopathic genes. Drosophila genetics provides more than 19,000 molecularly-defined P-elements inserted throughout the Drosophila genome that facilitate the generation of high-density genomic coverage. We have developed an innovative approach to phenotype cardiac function in adult awake Drosophila and have identified a P-element mutation in the short gastrulation (sog) gene that results in dilated cardiomyopathy. Based on our preliminary findings, we propose that mouse and Drosophila genetics can identify novel genes and mechanisms that are responsible for human dilated cardiomyopathies. Accordingly, we propose the following specific aims: Aim 1: To identify novel genes causing cardiomyopathy using an ENU phenotype-driven recessive screen in adult mice. We propose to map the disease causing gene located on chromosome 1 in the ENU family with abnormal cardiac function. Aim 2: To investigate the biochemical and genetic mechanisms through which mutations in the dpp/BMP signaling pathway lead to dilated cardiomyopathy in adult Drosophila. Genetic complementation experiments will be performed in Drosophila to prove that sog deficiency causes cardiomyopathy. Aim 3: To test whether decreased antagonism of the BMP pathway in mice, will result in a cardiomyopathic phenotype under conditions of pressure overload. TAG experiments will be performed in knock out mice deficient in the endogenous mammalian BMP antagonists chordin and noggin. Aim 4: To identify novel genes causing cardiomyopathy by performing a genome-wide screen of deletion mutants in Drosophila. Flies heterozygous for PiggyBac derived deletion mutants from the Exelixis collection will be screened for cardiomyopathy using optical coherence tomography followed by fine mapping of the disease-causing gene(s). Thus, these four integrated aims will harness the power of mouse and Drosophila genetics to identify and evaluate novel candidate genes for their role in cardiomyopathy.