The molecular basis of RECQ4-associated genetic disorders and cancer predispositi

RECQ4相关遗传性疾病和癌症易感性的分子基础

• 批准号: 8101561
• 负责人: Yilun Liu
• 金额: $ 8.4万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2011-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8101561
• 关键词: Aging-Related Process; Alleles; Amino Acids; Bacteria; Binding; Biochemical; CDC45L gene; Catalytic Domain; Cell Cycle; Cell Extracts; Cell Proliferation; Cell Survival; Cells; Chromatin; Chromosomes; Circadian Rhythms; Cisplatin; Complex; Cyclin-Dependent Kinases; DNA; DNA Binding; DNA Damage; DNA Repair; DNA Replication Factor; DNA Structure; DNA biosynthesis; Data; Defect; Development; Disease; Dissociation; Eukaryota; Evolution; Family; G1 Phase; Genes; Genome; Genomic Instability; Genomics; Genotoxic Stress; Goals; Health; Hereditary Disease; Homologous Gene; Human; In Vitro; Individual; Ionizing radiation; Knowledge; Lead; Lesion; Link; Lymphoma; MCM10 gene; MCM2 gene; Maintenance; Malignant Neoplasms; Maps; Metabolism; Methods; Molecular; Mutagenesis; Mutation; Organism; Outcome; Pathogenesis; Patients; Phenotype; Phosphorylation; Predisposition; Premature aging syndrome; Process; Prokaryotic Cells; Property; Protein Family; Proteins; RECQL4 gene; Regulation; Replication Initiation; Replication Origin; Reporting; Role; Set protein; Sister Chromatid; Tumor Suppressor Proteins; Vertebrates; Work; Yeasts; arm; base; biological adaptation to stress; cancer prevention; cancer therapy; cell growth; clinical phenotype; cohesion; helicase; in vitro activity; in vivo; metaplastic cell transformation; mouse model; osteosarcoma; polypeptide; prevent; protein complex; protein protein interaction; repaired; tumorigenesis

DESCRIPTION (provided by applicant): Functional machineries to prevent and repair DNA damages are required for maintaining genome integrity and preventing tumourigenesis. A set of proteins belonging to the RecQ family is among the cancer suppressors linked to repairing lesions caused by DNA damaging agents, such as ionizing radiation and cisplatin. During the course of evolution, RecQ genes appear to have been amplified and diverged from a single copy of the RecQ gene in bacteria and yeast to five RecQ homologs in humans. The clinical phenotypes caused by mutations in different RECQ proteins in humans and mouse models indicate that they have important and non-overlapping roles in maintaining genomic integrity and cancer prevention. Since RECQ helicases are highly interactive proteins and form large protein complexes in vivo, the unique functions of each of the RECQ proteins are likely defined and regulated by the specific protein-protein interactions they form in cells. To understand how the mammalian RECQ proteins act as tumor suppressors and caretakers for our genome, it is crucial to understand the unique functions of each of the RECQ protein complexes. Recently, our lab has several breakthroughs in the study of human RECQ4 helicase. First, we successfully reported ATP-dependent RECQ4 helicase activity in vitro. Surprisingly, domain analyses uncovered two distinct ATP binding and DNA unwinding activities within the RECQ4 protein. Secondly, we report the identification of a highly purified chromatin bound RECQ4 complex from human cell extracts. We found that essential replisome factors MCM10, MCM2-7 helicase, CDC45 and GINS are the primary interaction partner proteins of human RECQ4. Moreover, circadian proteins, TIMELESS and TIPIN important for cohesion establishment and DNA replication progression are part of the RECQ4 complex. Importantly, complex formation and the association of RECQ4 with the replication origin are cell cycle regulated. Our studies allow us to conclude that RECQ4 is an integral component of the MCM replicative helicase complex participating in DNA replication in human cells. Based on our work, the goal of this proposal is to define the exact function of RECQ4 complex in DNA replication, cohesion establishment and cancer avoidance. The Specific Aims are: (1) dissecting the role of human RECQ4 in DNA replication initiation; (2) Establishing the functional relationship between RECQ4 and TIM-TIPIN heterodimer in replication fork progression, genotoxic stress response and cohesion establishment; and (3) Analysis of the cancer-associated RECQ4 (c.1390+2delT) mutation. PUBLIC HEALTH RELEVANCE: DNA replication is essential for cell growth, and abnormal DNA synthesis can result in genome instability, mutagenesis and tumorigenesis. Therefore, elucidation of the mechanisms that regulate DNA replication and the molecular functions of the involved proteins, such as RECQ4, are fundamental to our understanding of the cancer development. Crucially, our detailed knowledge on the DNA replication factors will aid in the discovery of new cancer therapy treatments, since inhibiting DNA replication factors effectively prevents cell proliferation.

描述（由申请人提供）：需要预防和修复 DNA 损伤的功能性机制来维持基因组完整性和预防肿瘤发生。一组属于 RecQ 家族的蛋白质属于癌症抑制因子，与修复 DNA 损伤剂（例如电离辐射和顺铂）引起的损伤有关。在进化过程中，RecQ 基因似乎已被扩增，并从细菌和酵母中的 RecQ 基因的单个副本分化为人类中的五个 RecQ 同源物。人类和小鼠模型中不同 RECQ 蛋白突变引起的临床表型表明，它们在维持基因组完整性和癌症预防方面具有重要且不重叠的作用。由于 RECQ 解旋酶是高度相互作用的蛋白质并在体内形成大型蛋白质复合物，因此每个 RECQ 蛋白质的独特功能可能由它们在细胞中形成的特定蛋白质-蛋白质相互作用来定义和调节。为了了解哺乳动物 RECQ 蛋白如何充当我们基因组的肿瘤抑制因子和看护者，了解每个 RECQ 蛋白复合物的独特功能至关重要。 近期，我室在人RECQ4解旋酶研究方面取得多项突破。首先，我们在体外成功报道了 ATP 依赖性 RECQ4 解旋酶活性。令人惊讶的是，结构域分析发现了 RECQ4 蛋白内两种不同的 ATP 结合和 DNA 解旋活性。其次，我们报告了从人类细胞提取物中鉴定出高度纯化的染色质结合 RECQ4 复合物。我们发现必需的复制体因子MCM10、MCM2-7解旋酶、CDC45和GINS是人类RECQ4的主要相互作用伙伴蛋白。此外，对内聚力建立和 DNA 复制进程很重要的昼夜节律蛋白 TIMELESS 和 TIPIN 是 RECQ4 复合体的一部分。重要的是，复合物的形成以及 RECQ4 与复制起点的关联是细胞周期调节的。我们的研究使我们得出结论，RECQ4 是参与人类细胞 DNA 复制的 MCM 复制解旋酶复合体的一个组成部分。根据我们的工作，该提案的目标是确定 RECQ4 复合物在 DNA 复制、内聚力建立和癌症避免中的确切功能。具体目标是：（1）剖析人类RECQ4在DNA复制启动中的作用； (2) 建立RECQ4与TIM-TIPIN异二聚体在复制叉进展、基因毒性应激反应和内聚力建立中的功能关系； (3)癌症相关RECQ4 (c.1390+2delT)突变分析。 公共卫生相关性：DNA 复制对于细胞生长至关重要，DNA 合成异常可能导致基因组不稳定、突变和肿瘤发生。因此，阐明调节 DNA 复制的机制和相关蛋白质（如 RECQ4）的分子功能对于我们了解癌症发展至关重要。至关重要的是，我们对 DNA 复制因子的详细了解将有助于发现新的癌症治疗方法，因为抑制 DNA 复制因子可以有效防止细胞增殖。