Transposon dependent gene silencing in Drosophila

在包括人类在内的真核生物基因组中，存在大量的转座因子。这些短的DNA片段可以在基因组中移动，重新排列染色体的大片段，并改变基因的表达。它们在基因表达中的作用的遗传和分子机制还没有很好地理解，然而，很明显它们通过调节染色质结构的蛋白质发挥作用。对这些蛋白质及其功能的描述是理解所涉及机制的先决条件。为了理解这些机制，我们已经开发了一个简单的遗传系统，在常见的果蝇，果蝇，研究的作用，转座子，P元件，在相邻的报告基因的调节。果蝇是一个被证明的，优秀的模式生物。 从历史上看，遗传、细胞和发育生物学研究工具的优势为各种生物学和生化机制提供了宝贵的见解。我们过去的研究表明，P元件阻遏蛋白诱导基因沉默。其分子机制涉及特异性异染色质蛋白与DNA的结合。蛋白质-蛋白质和蛋白质-DNA相互作用的细节还不清楚。我们打算研究P元件阻遏物如何作用于引起位置特异性基因沉默的机制。使用已建立的方法，我们将把阻遏蛋白基因（和其他我们已经确定），在修改后的形式，到各种GAL 4-UAS表达构建体，并将它们转化回果蝇的种系，以评估其效果。我们假设，通过检查修饰（突变）形式的影响，我们可以确定这些蛋白质中的功能结构域，并推导出这种基因沉默的分子机制。 这些结构域可以通过其他方法如ChIP和免疫共沉淀来确认。蛋白质之间的结构域相互作用将定义构成该机制的分子事件。通过确定转座子介导的基因沉默在模式生物，如果蝇的分子机制，我们可以将这些知识应用到其他真核生物领域，如医学，农业和生物技术。

In eukaryote genomes, including humans, there are large numbers of transposable elements. These short DNA segments can move around the genome, rearrange large segments of chromosomes, and alter the expression of genes. The genetic and molecular mechanism(s) of their role in gene expression is not well understood, however, it is clear that they function via proteins that regulate chromatin structure. The description of these proteins and their functions is a prerequisite for understanding the mechanisms involved. Towards understanding these mechanisms we have developed a simple genetic system in the common Fruit Fly, Drosophila, to examine the role of a transposon, the P element, in the regulation of an adjacent reporter gene. Drosophila is a proven, excellent model organism. Historically, the advantages of the research tools in genetic, cellular, and developmental biology have provided valuable insight into a wide variety of biological and biochemical mechanisms. Our past research has shown that the P element repressor protein induces gene silencing. The molecular mechanism involves specific heterochromatic proteins binding to DNA. The details of the protein-protein and protein-DNA interactions are as yet unknown. We intend to examine the mechanism of how the P element repressor acts to cause location specific gene silencing. Using established methods, we will put the repressor protein gene (and others we have identified), in modified forms, into various GAL4-UAS expression constructs and transform them back into the germline of Drosophila to evaluate their effects. We hypothesize that by examining the effects of modified (mutant) forms we can identify functional domains in these proteins and derive the molecular mechanism(s) of this gene silencing. These domains can be confirmed by other methods such as ChIP and co-immunoprecipitation. The domain interactions between and among the proteins will define the molecular events constituting the mechanism. By identifying the molecular mechanism of transposon mediated gene silencing in a model organism, such as Drosophila, we can apply this knowledge to other eukaryotes in areas such as medicine, agriculture, and biotechnology.