Heterochromatin in the developing vertebrate embryo

描述(申请人提供)：异染色质是DNA的一种紧密包装形式，与转录抑制有关。它对正常发育和基因组稳定是必不可少的。异染色质异常与人类发育综合征免疫缺陷、着丝粒区域不稳定和面部异常(ICF)、退行性肌肉萎缩病FSHD(FSHD)有关，并可能促进癌症的非整倍体。异染色质是许多重要的生物学过程所必需的，包括正常的染色体分离、性染色体剂量补偿、印记基因的单等位基因表达和寄生DNA元件的转录沉默。到目前为止，我们对异染色质调控的大部分分子理解都来自于在植物、昆虫和真菌中进行的遗传筛选。从这些生物中筛选出的小鼠基因突变表明，异染色质调节的许多方面都是保守的，包括组蛋白3赖氨酸9甲基化和异染色质蛋白HP1的关键作用。然而，脊椎动物和其他模式生物的异染色质调控存在差异。例如，DNA修饰的5-甲基胞嘧啶是脊椎动物异染色质的重要成分，但不存在于裂殖酵母或黑腹果蝇的基因组中。斑马鱼提供了基因、分子和发育工具的强大组合，使研究脊椎动物异染色质的创新方法成为可能。这项建议使用这些工具来定义在重复序列中调节异染色质的分子通路，并确定正常发育和组织内稳所涉及的基因的需求。在斑马鱼异染色质调节剂的小屏幕上，我们的实验室鉴定了几个基因，包括zbtb24和nsd1a。ZBTB24在患有2型ICF综合征的人类中发生突变，NSD1在人类发育障碍Sotos综合征中发生突变。然而，编码蛋白在异染色质调节中的作用尚未在实验中得到解决。这项拟议的研究将定义依赖于这些因素的异染色质的内源性位点，并确定这些蛋白质在异染色质调节中的分子功能。在早期成功的基础上，将进行更广泛的候选和无偏见的功能丧失筛查，以识别和定义调节斑马鱼胚胎中异染色质的额外基因的分子功能。对这些基因的分子和发育需求的阐明有望揭示异染色质调节的新机制，这些机制在脊椎动物物种中是保守的，与人类发育和疾病状态相关。

DESCRIPTION (provided by applicant): Heterochromatin is a tightly packaged form of DNA that is associated with transcriptional repression. It is essential for normal development and genome stability. Abnormal heterochromatin is implicated in the pathology of the human developmental syndrome Immunodeficiency, Centromere region instability and Facial abnormalities (ICF), the degenerative muscle wasting disease Facioscapulohumeral muscular dystrophy (FSHD) and is likely to facilitate aneuploidy in cancer. Heterochromatin is required for a number of essential biological processes including normal chromosome segregation, sex chromosome dosage compensation, monoallelic expression of imprinted genes and transcriptional silencing of parasitic DNA elements. To date, much of our molecular understanding of heterochromatin regulation is derived from genetic screens performed in plants, insects and fungi. Mouse mutations in genes identified in screens from these organisms demonstrate conservation of many aspects of heterochromatin regulation, including critical roles for histone 3 lysine 9 methylation and the heterochromatin protein HP1. However, there are differences between heterochromatin regulation in vertebrates and other model organisms. For example, the DNA modification 5-methylcytosine is an essential component of heterochromatin in vertebrate species but is not present in the genomes of Schizosaccharomyces pombe or Drosophila melanogaster. Zebrafish offer a powerful combination of genetic, molecular and developmental tools that allow for innovative approaches to the study of vertebrate heterochromatin. This proposal uses these tools to define molecular pathways that regulate heterochromatin at repetitive sequences and to identify requirements for the involved genes in normal development and tissue homeostasis. In a small screen for regulators of zebrafish heterochromatin, our laboratory identified several genes including zbtb24 and nsd1a. ZBTB24 is mutated in humans with ICF syndrome type 2 and NSD1 is mutated in the human developmental disorder Sotos syndrome. However, roles for the encoded proteins in heterochromatin regulation have not been experimentally addressed. The proposed research will define endogenous sites of heterochromatin that depend on these factors and identify the molecular functions of these proteins in heterochromatin regulation. Based on early success, more extensive candidate and unbiased loss-of-function screens will be performed to identify and define the molecular functions of additional genes that regulate heterochromatin in the zebrafish embryo. Elucidation of the molecular and developmental requirements for these genes is expected to uncover novel mechanisms for heterochromatin regulation that are conserved in vertebrate species and which are relevant to human development and disease states.