Identification of DNA Elements Governing Chromatin Function in C elegans

描述(申请人提供)：真核基因组被包装成染色质，染色质调节蛋白质的功能，介导转录活动和其他基本过程，包括有丝分裂和减数分裂期间基因组的重组和忠实分离。这项建议的目标是在线虫中识别调节染色质结构和功能的离散元件，线虫是一种在大规模基因组研究和基因功能发现中具有核心重要性的模式后生动物。我们将首先使用ChlP-ChIP和相关方法来绘制选定的组蛋白修饰和染色体相关蛋白的基因组分布图，然后使用这些信息，结合来自其他modENCODE组的数据，建立染色质功能的定量模型。具体来说，我们会： 1.确定并从技术上验证控制染色质和染色体行为的功能元件。 我们分析的重点将是指定核小体的位置和占据、控制基因表达的区域、诱导X染色体的抑制、引导有丝分裂分离和基因组复制、控制减数分裂过程中的同源配对和重组以及组织染色体在核内的定位。126个战略选择的靶标包括关键的组蛋白修饰、组蛋白变体、RNA聚合酶II亚型、剂量补偿蛋白、着丝粒成分、同源配对促进剂、重组标记和核膜成分。高效的管道设计将有助于识别和验证与这些靶标相关的不同类别的功能元件，并将结果与注释良好的线虫基因组整合在一起。 2.对已鉴定的功能元件进行生物学验证，并建立完整的染色体功能定量模型。 我们将把Aim 1中产生的信息与现有的靶标生物学知识相结合，对缺乏选定靶蛋白的突变和RNAi提取物进行ChlP-Chip分析，使用染色体外阵列来评估候选已识别序列基序在体内招募靶标的能力，确定选定靶标的组织特异性模式，并创建转录和全染色体功能的集成、定量模型。 在线虫研究不断扩大和丰富的历史背景下实现这些目标，将为迎接利用基因组序列信息了解和预测生物功能的挑战提供一个重要的里程碑。

DESCRIPTION (provided by applicant): Eukaryotic genomes are packaged into chromatin, which regulates the function of proteins that mediate transcriptional activity and other essential processes, including recombination and the faithful segregation of the genome during mitosis and meiosis. The goal of this proposal is to identify discrete elements that regulate chromatin structure and function in the nematode C. elegans, a model metazoan of central importance in large-scale genomic research and gene function discovery. We will first use ChlP-chip and related methods to map the genomic distributions of selected histone modifications and chromosome-associated proteins, and then use that information, in combination with data from other modENCODE groups, to build quantitative models of chromatin function. Specifically, we will: 1. Identify and technically validate functional elements that control chromatin and chromosome behavior. The focus of our analysis will be elements that specify nucleosome positioning and occupancy, control domains of gene expression, induce repression of the X chromosome, guide mitotic segregation and genome duplication, govern homolog pairing and recombination during meiosis, and organize chromosome positioning within the nucleus. 126 strategically selected targets include key histone modifications, histone variants, RNA polymerase II isoforms, dosage-compensation proteins, centromere components, homolog-pairing facilitators, recombination markers, and nuclear-envelope constituents. An efficient pipeline design will facilitate identification and validation of the different classes of functional elements associated with these targets and will integrate the results with the well-annotated C. elegans genome. 2. Biologically validate identified functional elements and build integrated, quantitative models of chromosome function. We will integrate information generated in Aim 1 with existing knowledge on the biology of the targets, perform ChlP-chip analysis on mutant and RNAi extracts lacking selected target proteins, use extrachromosomal arrays to assess the ability of candidate identified sequence motifs to recruit targets in vivo, identify tissue-specific patterns of selected targets, and create integrated, quantitative models of transcription and whole-chromosome functions. Achieving these goals in the context of the ongoing expansion and rich history of C. elegans research will provide an important milestone in meeting the challenge of using genome sequence information to understand and predict biological functions.