Chromatin Biology of the African Trypanosome

非洲锥虫的染色质生物学

• 批准号: 10633288
• 负责人: Erik Debler
• 金额: $ 53.22万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-02 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10633288
• 关键词: Address; Affect; African; African Trypanosomiasis; Antigenic Variation; Area; Biochemical; Biological; Biological Assay; Biological Process; Biology; Cell Cycle; Cell Cycle Deregulation; Cell Cycle Progression; Cell Cycle Regulation; Cells; Chagas Disease; Chromatin; Complex; Cryoelectron Microscopy; DNA; DNA Binding; DNA biosynthesis; DNA replication fork; Data; Defense Mechanisms; Development; Disease; Enzymes; Epigenetic Process; Eukaryota; Exhibits; Flow Cytometry; Future; Gene Expression Regulation; Genetic; Genetic Transcription; Genomic DNA; Goals; Histone H3; Histones; Human; Hybrids; Immune system; In Vitro; Intervention; Kinetoplastida; Knowledge; Leishmania; Leishmaniasis; Life; Lysine; Medical; Methylation; Methyltransferase; Modeling; Modification; Molecular; Monitor; Mutagenesis; Nucleic Acid Regulatory Sequences; Nucleosome Core Particle; Nucleosomes; Organism; Parasites; Persons; Pharmaceutical Preparations; Ploidies; Populations at Risk; Process; Protozoa; RNA; RNA primers; Regulation; Research; Resistance; Roentgen Rays; Role; Signal Pathway; Structure; System; Testing; Therapeutic; Transcriptional Regulation; Trees; Trypanosoma; Trypanosoma brucei brucei; Trypanosoma cruzi; Variant; Work; X-Ray Crystallography; Yeasts; chromatin protein; combat; drug discovery; effective therapy; epigenetic regulation; experimental study; fungus; global health; histone methylation; in vivo; inducible gene expression; inhibitor; inhibitor therapy; insight; interest; mutant; neglected tropical diseases; new therapeutic target; novel; novel strategies; novel therapeutics; overexpression; pharmacologic; posttranscriptional; preference; reconstruction; recruit; spatiotemporal; structural determinants

PROJECT SUMMARY Protozoan parasites of the group kinetoplastids are responsible for major human maladies such as fatal sleeping sickness (Trypanosoma brucei, also termed the African trypanosome), Chagas disease (T. cruzi), and leishmaniasis (Leishmania species). Due to the lack of inexpensive and safe drugs, rising resistance against current drugs, and limited drug discovery efforts, novel approaches are urgently needed to combat these neglected tropical diseases. Because kinetoplastids constitute one of the earliest-branching organisms in the eukaryotic tree of life, they exhibit numerous molecular and cellular features that are distinct from metazoa and fungi, and that can be exploited for pharmacological intervention. By combining structural, biochemical, and in vivo approaches, we seek to address fundamental questions in chromatin biology and gene regulation in the model kinetoplastid T. brucei. We are particularly interested in the structure and mechanism of the closely related DOT1A and DOT1B enzymes that are key regulators of essential functions in T. brucei and that catalyze the methylation of histone H3 lysine 76 (H3K76) in the globular nucleosome core region. While DOT1A regulates cell-cycle progression, DOT1B in antigenic variation, an essential mechanism for the parasite to evade the host’s immune system. Due to significant mechanistic differences of trypanosome DOT1A/B to human and yeast DOT1 enzymes, the molecular mechanisms of how they methylate chromatin and how they are regulated remain poorly understood. In Aim 1, we will therefore investigate the mechanism of DOT1A-nucleosome substrate recognition and its impact on cell cycle control. In Aim 2, we will decipher the regulatory mechanism of DOT1A governed by RNaseH2, an enzyme that is known to cleave RNA in RNA/DNA hybrids and that has been implicated in both DNA replication and transcriptional regulation. Our goal is to define the impact of RNaseH2 on DOT1A activity, provide a structural basis for its regulatory function, and elucidate the mechanism of how DOT1A activity is coordinated with the cell cycle. The interaction of RNaseH2 with DOT1A/B is specific to trypanosomes, suggesting a novel regulatory mechanism of DOT1 enzymes. Collectively, our studies will illuminate the long- standing question of how DOT1A is recruited to chromatin and how it is regulated in a spatiotemporal manner. Our studies will yield the first atomic structures of the fundamental unit of chromatin, the nucleosome, of the vast group of protozoa, which are medically, ecologically, evolutionarily, and scientifically important eukaryotes. Due to the novel regulatory function of RNaseH2, our results will broaden our mechanistic understanding of DOT1A and RNaseH2 enzymes. Because T. brucei DOT1-regulated processes are essential for the parasite, this research may ultimately have a large impact on global health by exploiting the unique attributes of protozoan DOT1 structure and regulation to inform novel therapies for sleeping sickness and other diseases caused by kinetoplastids that affect half a billion of people.

项目概要 动质体的原生动物寄生虫是造成致命睡眠等主要人类疾病的原因 病（布氏锥虫，也称为非洲锥虫）、恰加斯病（T. cruzi）和 利什曼病（利什曼原虫属）。由于缺乏廉价且安全的药物，耐药性不断上升 由于现有的药物和有限的药物发现工作，迫切需要新的方法来应对这些问题 被忽视的热带疾病。因为动质体是自然界中最早分支的生物之一 真核生命树，它们表现出许多与后生动物不同的分子和细胞特征 真菌，可用于药理干预。通过结合结构、生物化学和 体内方法，我们寻求解决染色质生物学和基因调控的基本问题 模型动质体 T. brucei。我们对密切相关的结构和机制特别感兴趣 DOT1A 和 DOT1B 酶是 T. brucei 基本功能的关键调节因子，并催化 球状核小体核心区域组蛋白 H3 赖氨酸 76 (H3K76) 的甲基化。虽然 DOT1A 监管 细胞周期进程，抗原变异中的 DOT1B，是寄生虫逃避宿主免疫的重要机制 免疫系统。由于锥虫 DOT1A/B 与人类和酵母 DOT1 存在显着的机制差异 酶，它们如何甲基化染色质以及如何调节的分子机制仍然很差 明白了。因此，在目标 1 中，我们将研究 DOT1A-核小体底物识别的机制 及其对细胞周期控制的影响。在目标2中，我们将解密DOT1A的监管机制 RNaseH2，一种已知可切割 RNA/DNA 杂交体中 RNA 的酶，并且与这两种酶都有关联 DNA 复制和转录调控。我们的目标是确定 RNaseH2 对 DOT1A 活性的影响， 为其调节功能提供结构基础，并阐明DOT1A活性的机制 与细胞周期协调。 RNaseH2 与 DOT1A/B 的相互作用是锥虫特有的， 提出了 DOT1 酶的一种新的调控机制。总的来说，我们的研究将阐明长期 悬而未决的问题是 DOT1A 如何被招募到染色质以及它如何以时空方式进行调节。 我们的研究将产生染色质基本单位核小体的第一个原子结构 原生动物群，在医学、生态、进化和科学上都是重要的真核生物。到期的 RNaseH2 的新颖调节功能，我们的结果将拓宽我们对 DOT1A 机制的理解 和 RNaseH2 酶。由于 T. brucei DOT1 调节的过程对于寄生虫至关重要，因此 通过利用原生动物的独特属性，研究最终可能会对全球健康产生巨大影响 DOT1结构和调节为昏睡病和其他由昏睡病引起的疾病的新疗法提供信息 影响十亿人的动质体。