Mechanistic dissection of HP1 mediated heterochromatin

HP1介导的异染色质的机制剖析

• 批准号: 9060337
• 负责人: GEETA J NARLIKAR
• 金额: $ 42.78万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9060337
• 关键词: Address; Adopted; Affect; Behavior; Biochemical; Biological; Biological Models; Cells; Chromatin; Chromatin Structure; Chromosome Segregation; Complex; Cryoelectron Microscopy; DNA; Development; Dissection; Equilibrium; Fission Yeast; Foundations; Gene Activation; Gene Silencing; Genetic Recombination; Genetic Transcription; Genome; Goals; Health; Heterochromatin; Histone H3; In Vitro; Ligands; Link; Lysine; Malignant Neoplasms; Mediating; Metastatic breast cancer; Methods; Molecular; Molecular Conformation; Myeloid Leukemia; Nucleosomes; Play; Post-Translational Protein Processing; Process; Property; Protein Isoforms; Proteins; Recruitment Activity; Regulation; Role; Schizosaccharomyces pombe Proteins; Structure; Testing; Therapeutic Intervention; Work; analytical ultracentrifugation; base; chromatin protein; dimer; in vivo; in vivo Model; inhibitor/antagonist; leukemia; malignant breast neoplasm

DESCRIPTION (provided by applicant): The spread of heterochromatin is crucial for heritably silencing large regions of the genome and consequently for generating and maintaining cell identity during development. Illegitimate gene activation from loss of heterochromatin spread is strongly linked to invasive breast cancers while illegitimate gene silencing from aberrant heterochromatin spread is strongly linked to myeloid leukemias. In addition to gene silencing, heterochromatin plays crucial roles in recombination and chromosome segregation. At the core of the most conserved form of heterochromatin is the complex formed between HP1 proteins and chromatin methylated on lysine 9 of histone H3 (H3K9me3). The following key roles have been attributed to HP1 proteins in heterochromatin function: (1) The HP1-nucleosome complex is hypothesized to recruit effector molecules to H3K9me3 chromatin. Paradoxically, effectors that both, enable as well as restrict further heterochromatin spread are recruited. The balance between these opposing activities is thought to dictate the functions and stability of the assembled heterochromatin. (2) HP1 proteins are hypothesized to directly mediate the spread of heterochromatin by oligomerizing across multiple nucleosomes. (3) HP1 proteins are hypothesized to condense chromatin and thereby directly reduce the access of DNA to the transcription machinery. Despite the centrality of these properties to the in vivo functions of heterochromatin, the molecular basis for how HP1 accomplishes these roles is poorly understood. We have made the new discovery that Swi6, the major S. pombe HP1 isoform, switches from an auto-inhibited state to a spreading-competent state in a manner that depends on recognition of H3K9me3 and additional features of a nucleosome. We will build on these and additional results to address the following questions in the S. pombe model system: (i) how do HP1 proteins interact with different effectors, (iii) why do different HP1 isoforms have different functions, (iii) how do HP1 proteins spread across chromatin, and (iii) what does HP1 assembly do to chromatin structure? We will use a combination of quantitative biochemical methods and cutting edge electron cryo-microscopy approaches. We will also test key predictions of our models in vivo.

描述（由申请人提供）：异染色质的扩散对于基因组大区域的遗传沉默至关重要，从而在发育过程中产生和维持细胞身份。异染色质扩散丢失导致的非法基因激活与侵袭性乳腺癌密切相关，而异常异染色质扩散导致的非法基因沉默与骨髓性白血病密切相关。除了基因沉默之外，异染色质在重组和染色体分离中也起着至关重要的作用。最保守形式的异染色质的核心是 HP1 蛋白和组蛋白 H3 赖氨酸 9 (H3K9me3) 上甲基化的染色质之间形成的复合物。 HP1 蛋白在异染色质功能中具有以下关键作用：(1) 假设 HP1-核小体复合物将效应分子募集到 H3K9me3 染色质。矛盾的是，既能促进又能限制异染色质进一步扩散的效应器被招募。这些相反活性之间的平衡被认为决定了组装的异染色质的功能和稳定性。 (2) HP1 蛋白被假设通过跨多个核小体的寡聚来直接介导异染色质的扩散。 (3) HP1 蛋白被假设可以浓缩染色质，从而直接减少 DNA 与转录机器的接触。尽管这些特性对于异染色质的体内功能至关重要，但 HP1 如何实现这些作用的分子基础却知之甚少。我们有了新的发现，Swi6（主要的粟酒裂殖酵母 HP1 同工型）从自动抑制状态切换到有扩散能力的状态，其方式取决于对 H3K9me3 的识别和核小体的其他特征。我们将在这些结果和其他结果的基础上解决粟酒裂殖酵母模型系统中的以下问题：（i）HP1蛋白如何与不同的效应子相互作用，（iii）为什么不同的HP1亚型具有不同的功能，（iii）HP1蛋白如何在染色质上传播，以及（iii）HP1组装对染色质结构有何影响？我们将结合使用定量生化方法和尖端电子冷冻显微镜方法。我们还将在体内测试我们模型的关键预测。