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，每个细胞在六个小时内复制了30亿个核苷酸/碱基。一个 复杂且协调良好的蛋白质网络，协同工作以确保成功完成 过程。这些蛋白质的失活可能导致灾难性后果，例如基因组 不稳定性，癌症和许多其他使人衰弱的疾病。因此，任何十九种这样的缺陷 蛋白质导致一种称为范科尼贫血（FA）的毁灭性疾病。 FA是一种罕见的遗传疾病 以骨髓衰竭（BMF）的特征，血液学异常和非常高的发生率 恶性肿瘤。 BMF是FA患者死亡率的主要原因[67]和全年减少症，血小板减少症和 贫血在FA患者中很常见[68]。此外，足总患者的逐渐下降 造血茎和祖细胞细胞（HSPC）[67，69，70]，被认为是骨骼负责的 患者骨髓衰竭。但是，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。在过去的十年中，越来越多的 在重要的细胞途径中起作用的蛋白质正在分类为FA蛋白，突出显示 FA的复杂性。尽管FA蛋白的DNA修复介导的功能广泛与病因有关 在该疾病中，有驱动需要了解FA蛋白的整个细胞功能 外部交叉链接维修。最近的报告表明，FA途径在DNA复制中起了新作用。这 该提议的长期目标是研究有缺陷的复制，作为早期驱动力之一 Fanconi贫血患者的基因组不稳定性，导致造血干和祖细胞损耗。这 FA蛋白在DNA复制中的全基因组作用将通过研究它们的研究来更好地理解 参与促进基因组区域的复制，这些区域最容易受到复制应力的影响， 例如脆弱的站点。该提案的主要目标是阐明FA的机械参与 DNA复制中的蛋白质，使用脆弱位点作为模型基因组基因座。 Madireddy博士的初步研究， 证明FA蛋白，特别是FANCD2，即使在 没有外源复制应力。目标1的重点是识别结构元素，这可能是 摊位复制叉，在没有FA蛋白的情况下，并阐明了FANCD2的机制 减轻在CFS停顿的复制。了解为什么在没有的情况下改变了DNA复制启动 FANCD2蛋白AIM 2研究了FANCD2的染色质重塑作用。最后， 使用FA IPSC和重编程的造血干细胞（HSC）作为模型系统，AIM 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损耗 FA最早的阶段。我们希望这些提议的研究都大大增加了我们对 FA途径在DNA复制和基因组不稳定性中的机械参与，使我们能够 建立有关复制性缺陷对范科尼贫血病因的贡献的新范式。