Consequences of gain-of-function RPA1 mutations on telomere function and hematopoiesis

功能获得性 RPA1 突变对端粒功能和造血的影响

基本信息

批准号：
10704735
负责人：
Richa Sharma
金额：
$ 16.7万
依托单位：
ST. JUDE CHILDREN'S RESEARCH HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-15 至 2027-09-14
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10704735
关键词：
Affinity Area Award Binding Binding Proteins Biochemical Biological Biological Assay Biology Bone marrow failure CRISPR/Cas technology Cell Maintenance Cell Separation Cell division Cells Childhood Chromosomes Clinical Complex Cytogenetics DNA DNA Binding DNA Damage DNA Repair Disorder DNA Repair Gene DNA biosynthesis Data Development Plans Digestion Disease Disease model Educational workshop Ensure Exhibits Faculty Family Flow Cytometry Functional disorder Generations Genes Genetic Induction Genome Genome engineering Goals Hematopoiesis Hematopoietic Hematopoietic stem cells Heterogeneous-Nuclear Ribonucleoproteins Heterozygote Human Immunoprecipitation Impairment In Vitro Individual Inherited Institution K-Series Research Career Programs Length Liver Cirrhosis Location Maintenance Maps Mass Spectrum Analysis Mentors Methods Missense Mutation Molecular Mosaicism Multiprotein Complexes Mus Mutation Nuclear Proteins Organ Orthologous Gene Pathogenesis Pathologic Pathology Patient-Focused Outcomes Phenotype Physicians Process Prognosis Proteins Proteome Proteomics Publishing Pulmonary Fibrosis Research Research Support Resource Sharing Role S phase Saint Jude Children&apos s Research Hospital Scientist Single-Stranded DNA Stress System Telomerase Telomere Maintenance Telomere Shortening Testing Time Tissue imaging Variant animal resource career career development causal variant cellular imaging clinical care clinical phenotype experience gain of function gene therapy genetic pedigree imaging facilities improved in vivo induced pluripotent stem cell lectures light microscopy mouse model mutant novel nuclease premature preservation proband recruit repaired replication factor A replication factor C replication stress response segregation skills stem stem cell model stem cells success telomere

项目摘要

Project Summary Inherited mutations in genes governing telomere maintenance cause Telomere biology disorders (TBD), a group of diseases with a progressive course and poor prognosis characterized by pulmonary fibrosis, liver cirrhosis, and bone marrow failure (BMF). Understanding the molecular mechanisms governing telomere preservation and identifying TBD associated genes are established gaps. We recently published novel germline heterozygous missense variants in RPA1 in four unrelated probands presenting with TBD. We also demonstrated that RPA1 mutants have elevated affinity to random and telomere single strand DNA, which causes a unique gain-of- function (GoF) effect, defective hematopoiesis due to short telomeres in RPA1 mutant iPSC cells and extensive somatic rescue mosaicism. The molecular mechanism through which RPA1 GoF mutations cause telomere shortening, and TBD is unknown. I hypothesize that RPA1 mutations cause TBD by outcompeting essential telomere maintenance proteins for binding to ssDNA, thereby limiting stem cell replicative potential. The proposed studies will investigate these questions through two specific aims: 1) determine the molecular mechanism by which GoF RPA1E240K causes telomere dysfunction and 2) determine the effect of RPA1E240K on telomere maintenance and hematopoiesis in vivo. A Mentored Clinical Scientist Research Career Development Award (K08) will provide the candidate with the amount of protected time needed to achieve her career goal of independence as a physician scientist and focus on improving the clinical outcomes of patients with BMF and DNA repair disorders through understanding the mechanisms of their disease pathogenesis. A strong career development plan including experienced and successful faculty advisor and co-advisor committee, clear plans for progress assessments, and attendance at numerous courses, lectures, and workshops to increase proficiency in technical and management skills will accompany these research goals to ensure success as an independent physician scientist. This award will be completed at St. Jude Children’s Research Hospital (SJCRH), one of the world’s leading academic institutions focused on the research and treatment of pediatric catastrophic diseases, making it an exemplary location for an early career physician scientist to develop his career. In addition to the strong institutional support, St. Jude offers unmatched research support with facilities including the Flow Cytometry and Cell Sorting Shared Resource, Center for Advanced Genome Engineering (CAGE), Light Microscopy Division of Cell and Tissue Imaging Center, Animal Resource Center, Cytogenetics Core, and Center for Proteomics.

项目概要控制端粒维持的基因的遗传性突变导致端粒生物学疾病（TBD），一组以肺纤维化、肝硬化为特征的病程进行性和预后不良的疾病，了解控制端粒保存和骨髓衰竭的分子机制。 TBD 相关基因已确定缺口。我们最近发表了新的种系杂合子。四个不相关的先证者中存在 RPA1 错义变异，我们还证明了 RPA1。突变体对随机和端粒单链 DNA 的亲和力提高，这导致了独特的增益功能 (GoF) 效应、RPA1 突变 iPSC 细胞端粒短导致的造血缺陷以及广泛的 RPA1 GoF 突变引起端粒的分子机制。缩短，TBD 未知。我认为 RPA1 突变是通过竞争必需的来导致 TBD 的。端粒维持蛋白与单链DNA结合，限制干细胞的复制潜力。拟议的研究将通过两个具体目标来调查这些问题：1）确定分子 GoF RPA1E240K 导致端粒功能障碍的机制，2) 确定 RPA1E240K 对端粒功能障碍的影响体内端粒维持和造血作用。指导临床科学家研究职业发展奖（K08）将为候选人提供实现作为一名医师科学家的独立职业目标和专注所需的受保护时间通过了解 BMF 和 DNA 修复障碍患者的临床结果其疾病发病机制的强有力的职业发展计划，包括经验丰富的和成功的教师顾问和联合顾问委员会、明确的进度评估计划以及出席旨在提高技术和管理技能熟练程度的大量课程、讲座和讲习班将伴随这些研究目标，以确保作为一名独立医师科学家的成功。在世界领先学术机构之一的圣裘德儿童研究医院 (SJCRH) 完成专注于儿科重大疾病的研究和治疗，使其成为儿童重大疾病的典范。作为一名早期职业医师科学家，圣裘德的职业生涯发展除了强大的机构支持之外。通过流式细胞术和细胞分选共享等设施提供无与伦比的研究支持资源，高级基因组工程中心 (CAGE)，细胞和组织的光学显微镜部门成像中心、动物资源中心、细胞遗传学核心和蛋白质组学中心。