The multifaceted role of the Fanconi anemia tumor suppressor pathway in facilitating DNA replication

范可尼贫血肿瘤抑制途径在促进 DNA 复制中的多方面作用

• 批准号: 9755495
• 负责人: Advaitha Madireddy
• 金额: $ 24.9万
• 依托单位: RBHS -CANCER INSTITUTE OF NEW JERSEY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9755495
• 关键词: Abnormal Cell; Affect; Anemia; Automobile Driving; Binding; Biological Assay; Biological Models; Blood; Cell physiology; Cells; Cellular Stress; Chromosomal Breaks; Chromosomal Rearrangement; Chromosome Fragile Sites; Chromosome abnormality; Complement; Complex; Cytogenetics; DNA; DNA Damage; DNA Repair; DNA biosynthesis; DNA replication fork; Data; Defect; Disease; Elements; Ensure; Epigenetic Process; Etiology; Event; Fanconi Anemia pathway; Fanconi anemia protein; Fanconi' s Anemia; Fingers; Fishes; Fluorescent in Situ Hybridization; Future; Genetic Diseases; Genome; Genomic Instability; Genomic Segment; Genomics; Goals; Hematology; Hematopoietic; Hematopoietic stem cells; Histones; Hour; Human; Human Genome; Hybrids; Immunofluorescence Immunologic; Incidence; Knowledge; Lead; Light; Lymphocyte; Malignant Neoplasms; Maps; Mediating; Modeling; Molecular; Molecular Chaperones; Movement; Mutation; Nucleotides; Pancytopenia; Pathogenesis; Pathway interactions; Patients; Pattern; Physiological; Play; Predisposition; Process; Protein Deficiency; Proteins; RNA; Replication Initiation; Replication Origin; Reporting; Role; Site; Source; Stem cells; Stress; Structure; Syndrome; Thrombocytopenia; Time; Tumor Suppressor Proteins; Ubiquitin; Work; base; chromatin immunoprecipitation; chromatin modification; chromatin remodeling; crosslink; driving force; genome-wide; helicase; induced pluripotent stem cell; insight; interdisciplinary approach; mortality; nuclease; prevent; programs; protein function; rare genetic disorder; recessive genetic trait; recruit; repaired; replication stress; single molecule; tool

PROJECT SUMMARY During DNA replication, each cell copies three billion nucleotides/bases within a period of six-eight hours. A complex and well-coordinated network of proteins, work in tandem to ensure the successful completion of this process. The inactivation of any one these proteins, can lead to disastrous consequences like genomic instability, cancer and many other debilitating diseases. Accordingly, the deficiency in any one of nineteen such proteins results in a devastating disorder called Fanconi anemia (FA). FA is a rare genetic disorder characterized by bone marrow failure (BMF), hematological abnormalities and a very high incidence of malignancies. BMF is a leading cause of mortality in FA patients [67] and pancytopenia, thrombocytopenia and anemia are common in FA patients [68]. In addition, the FA patients have a progressive decline in hematopoietic stem and progenitors cells (HSPC) [67, 69, 70], which is thought be responsible for the bone marrow failure in patients. However, the primary mechanisms underlying HSPC decline, BMF and cancer predisposition in FA patients have remained elusive. Over the last decade, an increasing number of proteins functioning in important cellular pathways are being classified as FA proteins, highlighting the complexity of FA. While the DNA repair-mediated functions of FA proteins are widely implicated in the etiology of the disease, there is a driving need to understand the entire spectrum of cellular functions of FA proteins outside crosslink repair. Recent reports suggest a new role for the FA pathway in DNA replication. The long-term goal of this proposal is to investigate defective replication as one of the early driving forces of genomic instability, leading to hematopoietic stem and progenitor cell attrition, in Fanconi anemia patients. The genome-wide role of the FA proteins in DNA replication will be better understood by investigating their involvement facilitating the replication of regions of the genome that are most vulnerable to replication stress, such as fragile sites. The primary goal of this proposal is to elucidate the mechanistic involvement of the FA proteins in DNA replication, using fragile sites as model genomic loci. Preliminary studies by Dr. Madireddy, show that the FA proteins, specifically FANCD2, facilitates the replication of common fragile sites, even in the absence of exogenous replicative stress. The focus of Aim 1 is to identify structural elements, which potentially stall replication forks, in the absence of FA proteins and to elucidate the mechanism/s by which FANCD2 alleviates replication pausing at CFS. To understand why DNA replication initiation is altered, in the absence of the FANCD2 protein, Aim 2 investigates the chromatin remodeling role of FANCD2 is this process. Finally, using FA iPSC and reprogrammed hematopoietic stem cells (HSC) as model systems, Aim 3 investigates whether replication defects and the associated genomic instability are contributing to stem cell attrition in the earliest stages of FA. We expect that these proposed studies will both greatly increase our understanding of the mechanistic involvement of the FA pathway in DNA replication and genomic instability, and allow us to establish new paradigms regarding the contribution of replicative defects to the etiology of Fanconi anemia.

项目摘要 在DNA复制过程中，每个细胞在6 - 8小时内复制30亿个核苷酸/碱基。一 复杂和协调良好的蛋白质网络，协同工作，以确保成功完成这一任务。 过程这些蛋白质中任何一种的失活，都可能导致灾难性的后果，如基因组 不稳定，癌症和许多其他使人衰弱的疾病。因此，在任何一个十九个这样的缺陷， 这种蛋白质会导致一种叫做范可尼贫血（FA）的破坏性疾病。FA是一种罕见的遗传性疾病 其特征在于骨髓衰竭（BMF）、血液学异常和非常高的 恶性肿瘤。BMF是FA患者死亡的主要原因[67]，全血细胞减少症、血小板减少症和 贫血在FA患者中很常见[68]。此外，FA患者的 造血干细胞和祖细胞（HSPC）[67，69，70]，这被认为是负责骨 患者的骨髓衰竭。然而，HSPC下降、BMF和癌症的主要机制 FA患者的易感性仍然难以捉摸。在过去的十年里，越来越多的 在重要的细胞通路中起作用的蛋白质被归类为FA蛋白质，突出了 FA的复杂性。虽然FA蛋白的DNA修复介导的功能在病因学中广泛涉及 对于这种疾病，迫切需要了解FA蛋白的整个细胞功能谱 外部交联修复。最近的报道表明FA途径在DNA复制中的新作用。的 本提案的长期目标是研究缺陷复制作为早期驱动力之一， Fanconi贫血患者的基因组不稳定性，导致造血干细胞和祖细胞磨损。的 FA蛋白在DNA复制中的全基因组作用将通过研究其 参与促进最易受复制应激影响的基因组区域的复制， 例如脆弱的网站。这项提案的主要目的是阐明足总的参与机制 DNA复制中的蛋白质，使用脆性位点作为模型基因组位点。马迪雷迪博士的初步研究 显示FA蛋白，特别是FANCD 2，促进了常见脆性位点的复制，即使在 缺乏外源性复制应激。目标1的重点是确定结构要素， 停止复制叉，在FA蛋白的情况下，并阐明FANCD 2 在CFS处暂停复制。为了理解为什么DNA复制起始被改变， FANCD 2蛋白，目的2探讨FANCD 2在染色质重塑中的作用。最后， 使用FA iPSC和重编程造血干细胞（HSC）作为模型系统，目的3研究 复制缺陷和相关的基因组不稳定性是否有助于干细胞的磨损， 最早的阶段FA我们希望这些拟议的研究将大大增加我们对 FA途径在DNA复制和基因组不稳定性中的机制参与，并允许我们 建立关于复制缺陷对范可尼贫血病因学的贡献的新范式。