Genome-wide profiling of histone variants in Drosophila and Caenorhabditis

描述（由申请人提供）： 我们建议以高分辨率绘制整个注释的果蝇和秀丽隐杆线虫基因组的主要核心组蛋白变体。这些图谱将提供一个基于染色质的框架，用于解释由他人确定的表观遗传信息和基因表达模式。为了实现这一点，我们将使用生物素标记系统，我们已经成功地应用于基因组规模的染色质亲和纯化与标准的微阵列读出平台。在两个D. melanogaster和C.我们将分析通用的非复制依赖性组蛋白3变体H3.3，以及对照复制偶联变体H3。这些数据将提供两种模式生物中组蛋白动态的高分辨率全基因组图谱。H3.3模式与全基因组范围内的活性组蛋白修饰模式以及同源异型基因簇处的DNase1超敏位点的密切对应关系表明，我们的全基因组图谱可用于验证旨在绘制这些其他基因组特征的努力。在两个D. melanogaster和C.我们还将分析H2AZ变体，该变体与各种不同生物体中的表观遗传过程有关。果蝇H2AZ作为H2AX变体（称为H2AV）发挥着独特的双重作用，我们的初步研究表明，H2AV的模式与H3.3不同，从而提供了全基因组蛋白动力学的不同表观遗传特征。为了扩大我们对这两类组蛋白变体功能作用的理解，我们将使用RNAi敲除果蝇细胞系和整个C.在不同的发育阶段。敲低变体及其组装机器后的转录谱分析将提供它们在基因调控中所起作用的功能验证，并且敲低后的组蛋白变体谱分析应有助于鉴定靶基因。与其他类型的全基因组信息一起，我们的组蛋白变异图谱可能有助于更全面地了解这两种模式生物基因组中的表观遗传调控元件。考虑到相同的组蛋白变体被认为在基本上所有复杂的真核生物中起着相似的作用，我们的基本方法可以立即应用于人类基因组。

DESCRIPTION (provided by the applicant): We propose to map major core histone variants throughout the annotated Drosophila melanogaster and Caenorhabditis elegans genomes at high resolution. These maps will provide a chromatin-based framework for interpreting epigenetic information and gene expression patterns determined by others. To accomplish this, we will use the biotin-tagging system that we have successfully applied for genome-scale chromatin affinity purification together with a standard microarray readout platform. In both D. melanogaster and C. elegans we will profile the universal replication-independent histone 3 variant, H3.3, as well as the control replication-coupled variant, H3. These data will provide high-resolution genome-wide maps of histone dynamics in both model organisms. The close correspondences of H3.3 patterns to patterns of active histone modifications genome-wide and with DNasel hypersensitive sites at homeotic gene clusters suggests that our genome-wide maps can be used to validate efforts aimed at mapping these other genomic features. In both D. melanogaster and C. elegans we will also profile the H2AZ variant, which has been linked to epigenetic processes in a variety of different organisms. Drosophila H2AZ plays a unique dual role as the H2AX variant, called H2AV, and our preliminary studies indicate that patterns of H2AV differ from those of H3.3, thus providing a different epigenetic profile of histone dynamics genome-wide. To expand our understanding of the functional roles of both classes of histone variants, we will use RNAi to knock down the function of key chromatin regulators in Drosophila cell lines and in whole C. elegans at different developmental stages. Transcriptional profiling after knock-down of the variants and their assembly machines will provide functional validation of roles that they play in gene regulation, and histone variant profiling after knock-down should help to identify target genes. Together with other types of genome-wide information our histone variant maps are likely to contribute to a fuller understanding of epigenetic regulatory elements in the genomes of these two model organisms. Given that the same histone variants are thought to play similar roles in essentially all complex eukaryotes, our basic approach can be immediately applied to the human genome.