Drosophila as a Model to Investigate Rett Pathogenesis

描述（由申请人提供）：编码甲基cpg结合蛋白2 （MeCP2）的基因突变导致Rett综合征（RTT）和许多其他男性和女性的智力迟钝综合征。RTT是由MECP2的功能缺失突变引起的，MECP2结合靶DNA并招募Sin3A和组蛋白去乙酰化酶来沉默转录。MeCP2是一个更大的基因家族的一部分，甲基cpg结合域（MBD）家族，从果蝇到人类都是保守的。在果蝇中，使用UAS-GAL4系统过度表达MECP2会产生一种表型，可能是通过与MBD基因竞争产生的。我建议继续对全长系进行表征，并创建额外的转基因果蝇：两个系过度表达在Rett患者中常见的MECP2突变，一个系产生不含MBD区域的MECP2。其次，我将表征果蝇MBD基因CG10042的天然RNA和蛋白质表达模式。然后，我将通过p元素诱变和基因组eDNA杂交的RNAi来研究它的功能。我将描述所有产生的表型，并使用这些菌株通过f1修饰剂筛选来鉴定MeCP2和/或CG10042的修饰剂，以深入了解MeCP2功能的途径，迄今为止仅在体外进行了研究。通过提出的基因研究，我希望能够确定蛋白质，然后研究Rett和相关疾病的精神和神经功能障碍的机制。

DESCRIPTION (provided by applicant): Mutations in the gene encoding methyl-CpG-binding protein 2 (MeCP2) cause Rett syndrome (RTT) and a host of other mental retardation syndromes in both males and females. RTT results from loss-of-function mutations of MECP2, which binds to target DNA and recruits Sin3A and histone deacetylases to silence transcription. MeCP2 is part of a larger family of genes, the methyl-CpG-binding domain (MBD) family, which is conserved from fly through humans. Overexpression of MECP2 in Drosophila using the UAS-GAL4 system produces a phenotype, possibly by competing with MBD genes. I propose to continue characterization of the full-length lines and create additional transgenic flies: two lines which overexpress MECP2 mutations commonly found in Rett patients and one line which produces MeCP2 without the MBD region. Secondly, I will characterize the Drosophila MBD gene CG10042 for native RNA and protein expression patterns. Then I will investigate its function through P-element mutagenesis and RNAi with genomic eDNA hybrids. I will characterize all resulting phenotypes and use these strains to identify modifiers of MeCP2 and/or CG10042 through an F 1 modifier screen to gain insight into pathways of MeCP2 function, which until now has been studied only in vitro. Through the proposed genetic studies, I hope to identify proteins that can be then studied to investigate the mechanisms of the mental and neurological dysfunction in Rett and related disorders.