The role of the nucleolus in human genome organization in normal and disease states

正常和疾病状态下核仁在人类基因组组织中的作用

• 批准号: 10117559
• 负责人: Daniel Richard Foltz
• 金额: $ 64.3万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-21 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10117559
• 关键词: 3-Dimensional; Affect; Architecture; Biology; Cancer Biology; Cell Nucleolus; Cell Nucleus; Cell Wall; Cells; Centromere; Characteristics; Chromatin; Chromosome Segregation; Chromosomes; Cis-Acting Sequence; Clinical Trials; Complex; Cytology; DNA; Data; Data Set; Disease; Eukaryota; Event; Frequencies; Gene Dosage; Gene Expression; Genetic Transcription; Genome; Genome Stability; Genomics; Goals; Health; Heterochromatin; Histones; Human; Human Chromosomes; Human Genome; Immune; Immune signaling; Infection; Inflammatory; Innate Immune System; Interphase Cell; Investigation; Kinetics; Label; Learning; Link; Lipopolysaccharides; MAP Kinase Gene; Malignant Neoplasms; Maps; Measurement; Mediating; Methods; Mitotic spindle; Modification; Molecular; Mus; Natural Immunity; Neoplasm Metastasis; Normal Cell; Nuclear; Nucleolar Proteins; Organelles; Phase; Prevalence; Process; Property; Proteins; RNA; RNA-Binding Proteins; Repetitive Sequence; Research Personnel; Ribosomal RNA; Ribosomes; Role; Satellite DNA; Series; Shapes; Signal Pathway; Signal Transduction; Site; Stimulus; Stress; Structure; Surface; System; Testing; Therapeutic; Transducers; Untranslated RNA; anticancer treatment; base; cancer cell; carcinogenesis; carcinogenicity; cell type; cellular targeting; chromatin modification; chromosome missegregation; chromosome movement; deep sequencing; experimental study; extracellular; functional outcomes; genome-wide; genomic locus; histone modification; macrophage; malignant phenotype; member; monocyte; neoplastic cell; novel; p38 Mitogen Activated Protein Kinase; pathogen; recruit; response; therapeutic lead compound; tool; tumor xenograft

7. Project Summary / Abstract In all eukaryotes, the largest nuclear body is the nucleolus, a phase-separated, non-membrane bound organelle specialized for the synthesis of ribosomal RNAs and their assembly into ribosomes. Additionally, the exterior of the nucleolus is a hub for interactions with multiple specific DNA loci, thereby contributing to the three-dimensional architecture of the eukaryotic nucleus. Nucleolus-genome interactions are intimately connected to processes central to human health. For example, nucleolar-associated DNA is highly enriched in centromeric repetitive sequences. Centromeres, the sites of chromosome attachment to mitotic spindles, are fundamentally important for proper chromosome segregation. Several nucleolar proteins have been implicated in centromere-nucleolar interactions, and several centromeric proteins prominently reside in nucleoli in interphase cells. We have found that the nucleolar- centromeric interactions are regulated during cellular differentiation and are greatly increased in cancer cells. However, the mechanisms that regulated these interactions remain unknown. Not only do cancer cells display increased centromere-nucleolar interactions, they also frequently contain a perinucleolar compartments (PNC), a complex cytological feature that is absent in non-tumor cells. PNCs are located on the surface of nucleoli and contain multiple RNA species and RNA-binding proteins. We demonstrate here that these bodies also contain specific DNA loci, some of which encode non-coding RNAs retained within PNCs. A candidate cancer therapeutic termed metarrestin was isolated based on its ability to dissociate PNCs; metarrestin is currently in clinical trials based on its ability to reduce metastasis in human tumor xenograft experiments. Importantly for this proposal, we have observed that metarrestin also perturbs centromere-nucleolar interactions. We also present data that centromere-nucleolus interactions are perturbed in macrophages upon exposure the bacterial lipopolysaccharide (LPS), a canonical stimulus for the innate immune system. We also show that this response is blocked upon inhibition of specific signaling pathways. These changes are accompanied by altered nuclear distribution of the H3K27me3, a histone modification characteristic of facultative heterochromatin. Altogether, the central theme of this proposal is that the factors that govern centromere-nucleolus interactions are important for understanding chromosome missegregation, metastasis, and innate immunity. We plan a series of synergistic experiments to learn more about the underlying mechanisms. For example, we will test whether the centromeric activity of neocentromeres generates nucleolar associations, or if instead that is a property of centromeric satellite repeats regardless of activity. We will take candidate and unbiased approaches to finding centromeric proteins required for nucleolar interactions. We will characterize how metarrestin affects association of DNA loci with PNCs and nucleoli, and we will define cis-acting loci involved in PNC association. We will characterize the signaling pathways required for signaling-mediated disruption of nucleolar-centromeric interactions in macrophages. Results from these studies will allow for subsequent testing of universality. For example, do signaling components in macrophages also operate in tumor cells when treated with the therapeutic metarrestin? In this manner, this collaborative proposal will unite questions from diverse experimental systems to answer questions about the fundamental links between nuclear organization and human health.

7。项目摘要 /摘要 在所有真核生物中，最大的核体是核仁，一个相位分离的非膜结合 细胞器专门用于合成核糖体RNA及其组装成核糖体。另外， 核仁的外部是与多个特定DNA基因座相互作用的枢纽，从而有助于 真核核的三维结构。 核酚基因组相互作用与人类健康中心的过程密切相关。为了 例如，与核仁相关的DNA高度富集在丝粒重复序列中。 Centromeres， 染色体附着在有丝分裂纺锤体上的位点，对于适当的染色体至关重要 隔离。几种核仁蛋白已与中心粒核相互作用有关，几个 丝粒蛋白突出存在于相间细胞中的核仁中。我们发现核仁 - 在细胞分化过程中，丝粒相互作用受到调节，并且在癌细胞中大大增加。 但是，调节这些相互作用的机制仍然未知。 癌细胞不仅显示出增加的中心粒核相互作用，而且还经常包含 核酸室室（PNC），这是一种复杂的细胞学特征，在非肿瘤细胞中不存在。 PNC是 位于核仁的表面上，包含多种RNA物种和RNA结合蛋白。我们 在这里证明这些物体还包含特定的DNA基因座，其中一些编码非编码RNA 保留在PNC中。候选癌症治疗所谓的Metarrestin是根据其的能力隔离的 解离PNC； Metarrestin目前正在临床试验中，基于其减少人类转移的能力 肿瘤异种移植实验。重要的是，对于此提案，我们观察到Metarrestin也 Centromere核酸相互作用。 我们还提供了数据，表明在暴露后，巨噬细胞在巨噬细胞中互动 细菌脂多糖（LPS），一种天生免疫系统的规范刺激。我们还表明 抑制特定信号通路后，这种反应被阻止。这些变化伴随着 H3K27me3的核分布改变，这是一种组蛋白修饰的特征 异染色质。 总的来说，该提案的中心主题是管理着丝粒核糖的因素 相互作用对于理解染色体的错误分析，转移和先天免疫力很重要。 我们计划一系列协同实验，以了解有关潜在机制的更多信息。例如，我们 将测试新中心粒子的cencer粒活性是否会产生核酸关联，还是 无论活动如何，无论活动如何我们将服用候选人和公正 寻找核仁相互作用所需的丝粒蛋白的方法。我们将表征如何 Metarrestin会影响DNA基因座与PNC和核仁的关联，我们将定义与涉及的顺式作用基因座 PNC协会。我们将表征信号介导的破坏所需的信号传导途径 巨噬细胞中的核仁中心相互作用。这些研究的结果将允许随后的测试 普遍性。例如，当巨噬细胞中的信号传导成分在肿瘤细胞中也有效 用治疗性玛甲治疗？通过这种方式，该协作提议将团结起来 各种实验系统回答有关核组织之间基本联系的问题 和人类健康。