Chemical Strategies to Investigate Gene Regulation by Histone SUMOylation

研究组蛋白 SUMO 化基因调控的化学策略

• 批准号: 9548772
• 负责人: Champak Chatterjee
• 金额: $ 5.61万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-05 至 2019-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9548772
• 关键词: Acetylation; Acute; Address; Adopted; Affect; Antibodies; Biochemical; Biochemistry; Biological Assay; Biology; Biophysics; Brain; Cell Nucleus; Cells; Cellular biology; Chemicals; Chromatin; Chromatin Modeling; Chromatin Remodeling Factor; Chromatin Structure; Chromosomes; Complex; Cultured Cells; DNA; DNA Double Strand Break; Disease; Disease Progression; Enzymes; Euchromatin; Eukaryota; Fragile X Syndrome; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Goals; Growth; Heart Hypertrophy; Hematopoietic Neoplasms; Heterochromatin; Histone Acetylation; Histone Deacetylase; Histone H2B; Histone H4; Histones; Human; Human Cell Line; In Vitro; Knowledge; Laboratories; Lead; Ligase; Link; Lung; Lysine; Malignant Neoplasms; Malignant neoplasm of brain; Mediating; Methodology; Methods; Modification; Molecular; Molecular Biology; Multienzyme Complexes; Names; Nucleosomes; Organic Chemistry; Pattern; Physiological; Post-Translational Protein Processing; Process; Proteins; Proteomics; Regulation; Regulatory Pathway; Renal carcinoma; Research; Role; Rubinstein-Taybi Syndrome; SECTM1 gene; Series; Side; Signaling Protein; Site; Structure; Testing; Therapeutic; Therapeutic Intervention; Transcription Elongation; Ubiquitin; Yeasts; base; biophysical analysis; chemical group; chromatin modification; chromatin remodeling; experimental study; functional outcomes; gene function; gene repair; gene repression; genome-wide; heterochromatin-specific nonhistone chromosomal protein HP-1; histone demethylase; histone modification; human disease; in vitro Assay; in vivo; invention; malignant breast neoplasm; new therapeutic target; overexpression; prevent; programs; protein complex; protein protein interaction; public health relevance; reconstitution; repaired; segregation; tool; transcription factor

DESCRIPTION (provided by applicant): The long-term goal of the research in this proposal is to gain a molecular understanding of chromatin regulation by histone modification with the small ubiquitin-like modifier (SUMO) protein. The post-translational modification (PTM) of histone proteins by a range of chemical groups is observed in all eukaryotes. An extensive body of work has established that the dynamic regulation of histone PTMs and the biochemical relationships between specific PTMs underlie critical processes such as DNA transcription, repair, and replication. One dramatic PTM, the conjugation of histone lysine side-chains with the protein SUMO (termed SUMOylation) occurs widely, from yeast to humans, and is implicated in transcriptional silencing and DNA double-strand break repair. The dysregulation of these critical processes by environmental or genetic factors is linked to many human diseases such as cancers of the blood, brain, breast, and kidneys, to name a few. Therefore elucidating the molecular mechanisms by which histone SUMOylation regulates transcription and gene repair are essential first steps toward devising rational therapeutic strategies for acute human diseases. The biophysical and biochemical characterization of SUMOylated chromatin has until recently been limited by the inability to obtain sufficient quantities of homogeneously SUMOylated histones for in vitro studies, either from cultured cells or by enzymatic means. Hence, essentially nothing is known about the direct and/or indirect mechanisms by which histone SUMOylation influences the structure and function of human chromatin. In order to address this significant gap in our knowledge, we aim to combine the tools of synthetic organic chemistry, biochemistry, molecular and cell biology. Our chemical biology-based approach involves synthesizing site-specifically SUMOylated histones for biochemical and biophysical studies, as well as generating antibodies to investigate the genome-wide occurrence of SUMOylated histones. By adopting methods to study histone SUMOylation both in vitro and in cells, we will gain a comprehensive understanding of the mechanistic roles for this modification in normal growth and in disease progression. The specific research objectives of this proposal are: (1) To elucidate the direct effects of SUMOylation on chromatin structure and stability. (2) To elucidate the biochemical relationship between histone SUMOylation and gene-activating histone modifications, and (3) To investigate the histone SUMOylation state of active and silent regions of human chromatin. The successful completion of these aims will lead to a molecular understanding of SUMO-mediated changes in chromatin structure and function. Identifying new protein- protein interactions in regulatory pathways involving SUMOylated histones may provide new targets for therapeutic invention in diseases arising from the dysregulation of histone modifications. Finally, the methodologies described in this proposal are broadly applicable and will serve as transformative tools for studying SUMO-mediated processes in the context of other key signaling proteins, such as transcription factors, DNA- and histone-modifying enzymes.

描述（由申请人提供）：该提案中研究的长期目标是通过使用小型泛素样修饰剂（SUMO）蛋白质修饰组蛋白来获得对染色质调节的分子理解。在所有真核生物中都观察到了一系列化学基团对组蛋白的翻译后修饰（PTM）。广泛的工作已经确定，组蛋白PTM的动态调节以及特定PTM之间的生化关系是临界过程，例如DNA转录，修复和复制。一个戏剧性的PTM，是组蛋白赖氨酸侧链与蛋白质相扑（称为sumoylation）的结合，从酵母到人都广泛发生，并且与转录沉默和DNA双链破裂修复有关。通过环境或遗传因素对这些关键过程的失调与许多人类疾病（如血液，大脑，乳腺癌和肾脏的癌症）有关，仅举几例。因此，阐明组蛋白Sumoylation调节转录和基因修复的分子机制是为急性人类疾病制定理性治疗策略的重要第一步。直到最近才能从培养细胞或酶促手段获得足够数量的同质共酰基化组蛋白来获得足够数量的均合并组蛋白进行体外研究，直到最近才能限制Sumoypal染色质的生物物理和生化表征。因此，从本质上讲，对组蛋白sumoylation影响人类染色质的结构和功能的直接和/或间接机制一无所知。为了解决我们的知识这一显着差距，我们旨在结合合成有机化学，生物化学，分子和细胞生物学的工具。我们的基于化学生物学的方法涉及用于生化和生物物理学研究的特定于特定于特异性的组合组蛋白，以及生成抗体，以研究范围全基因组的Sumoymet组蛋白的发生。通过在体外和细胞中采用研究组蛋白企业的方法，我们将对这种修饰在正常生长和疾病进展中的机理作用有全面的理解。该提案的具体研究目标是：（1）阐明sumoylation对染色质结构和稳定性的直接影响。 （2）阐明组蛋白sumoylation和基因激活组蛋白修饰之间的生化关系，以及（3）研究人染色质的活性和沉默区域的组蛋白sumoylation态。 这些目标的成功完成将导致对Sumo介导的染色质结构和功能变化的分子理解。鉴定涉及sumoy酰基组蛋白的调节途径中的新蛋白质相互作用可能为因组蛋白修饰的失调而引起的疾病的治疗发明提供新的靶标。最后，本提案中描述的方法非常适用，并将作为在其他关键信号蛋白（例如转录因子，DNA和组蛋白修饰酶）背景下研究SUMO介导的过程的变革工具。