MOLECULAR CONSEQUENCES OF TELOMERASE DYSFUNCTION DURING HEMATOPOIETIC DEVELOPMENT

造血发育过程中端粒酶功能障碍的分子后果

基本信息

批准号：
9363584
负责人：
Luis Francisco Zirnberger Batista
金额：
$ 38.13万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-01 至 2022-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9363584
关键词：
Adult Affect Age Aplastic Anemia CRISPR/Cas technology Cell Differentiation process Cell Survival Cell physiology Cells ChIP-seq Clinical Complex DNA Damage Data Degradation Pathway Development Disease Dyskeratosis Congenita Engineering Event Failure Functional disorder Gene Expression Gene Expression Regulation Genes Genetic Genome engineering Goals Hematopoiesis Hematopoietic Homeostasis Human Impairment In Vitro Inherited Knowledge Length Maintenance Mammalian Cell Mediating Methods MicroRNAs Modeling Molecular Mutation Output Pancytopenia Pathogenesis Pathway interactions Patient-Focused Outcomes Patients Pattern Phosphodiesterase I Pluripotent Stem Cells Population Positioning Attribute Protocols documentation RNA Decay RNA Degradation RNA Interference RNA-Directed DNA Polymerase Regulation Research Rest Ribonucleoproteins Role Signal Transduction Source Specific qualifier value Stem cells Syndrome System TP53 gene Telomerase Telomerase RNA Component Telomere Maintenance Telomere Shortening Treatment outcome biochemical tools bone marrow failure syndrome cell type cytopenia disease phenotype exosome experimental study human disease human embryonic stem cell human pluripotent stem cell improved knock-down mutant novel patient population programs response stem cell biology telomere therapy development tool

项目摘要

PROJECT SUMMARY/ABSTRACT The overarching theme of our research is to decipher the molecular events responsible for bone marrow failure (BMF) in patients with impaired telomere maintenance, with the goal of improving treatment outcomes in these patients. The focus of this proposal is to use human pluripotent stem cells (hPSCs) as a novel platform to understand the molecular regulation of definitive hematopoietic impairment in cells with disease-associated mutations in telomerase, and to find novel alternatives to increase hematopoietic output in cells with dysfunctional telomeres. Dyskeratosis congenita (DC) is an inherited BMF syndrome where patients have very short telomeres for their age, typically below the first percentile length when compared to the rest of the population. All mutations discovered in DC are in genes that affect telomere homeostasis, including mutations in the telomerase components TERT, TERC, and dyskerin (DKC1). Difficulties in isolating relevant cell populations from DC patients have precluded the development of therapies against this disease. We recently overcame this limitation and combined genome engineering in hPSCs with in vitro human hematopoietic differentiation methods to create a novel and robust platform to study the mechanisms of hematopoietic failure caused by telomere dysfunction. In this submission, three specific aims are proposed that utilize this system to understand the molecular consequences of telomerase impairment during hematopoietic development, and to decipher novel targets to rescue hematopoietic output in cells harboring eroded telomeres. In aim 1, we will decipher the role of DNA damage responses (DDR) in the progression of definitive hematopoietic failure from cells with low levels of TERC. In particular, we will focus on the role of the p53 pathway in the hematopoietic impairment of cells with short telomeres, decipher the roles of miR-34a and miR-145 during abnormal hematopoiesis and decipher the genetic consequences of silencing DDR to restore hematopoietic output in cells harboring DC- associated mutations. In aim two we will decipher if the modulation of RNA decay in cells harboring mutations in DKC1 is a viable strategy to restore hematopoietic output in these cells. We will modulate TERC posttranscriptional processing and degradation by inhibition of specific RNA decay pathways. Finally, in aim 3, we will investigate if novel, non-canonical roles of TERC outside telomere maintenance can influence hematopoietic output and potentiate disease phenotypes in patients harboring mutations that reduce TERC levels, since these are commonly afflicted with severe disease phenotypes. Collectively, the studies proposed will allow us to decipher novel targets for directed therapies in patients suffering with telomere-syndromes. Our unique cellular and molecular tools, combined with our expertise in telomerase, stem cell biology and hematopoiesis puts us in an ideal position to make a significant impact in this field.

项目摘要/摘要我们研究的总体主题是破译负责骨骼的分子事件端粒维持受损患者的骨髓衰竭（BMF），目的是改善治疗这些患者的结果。该建议的重点是将人类多能干细胞（HPSC）用作了解细胞中明确造血损伤的分子调节的新型平台端粒酶中与疾病相关的突变，并找到新的替代方法，以增加造血产量端粒功能失调的细胞。兴奋性兴奋性葡萄球菌（DC）是一种遗传性的BMF综合征，患者的端粒非常短对于他们的年龄，与其他人口相比，通常低于第一个百分位长度。全部在DC中发现的突变是影响端粒稳态的基因，包括突变端粒酶成分TERT，TERC和DYSKERIN（DKC1）。隔离相关细胞种群的困难来自DC患者的患者排除了针对该疾病的疗法的发展。我们最近克服了这种限制和与体外人造血分化的HPSC中的基因组工程合并创建一个新颖而健壮的平台的方法，以研究由端粒功能障碍。在此提交中，提出了三个具体目标，以利用该系统来了解造血发育过程中端粒酶损伤的分子后果，并破译新型拯救具有侵蚀端粒侵蚀的细胞中造血输出的靶标。在AIM 1中，我们将破译角色 DNA损伤反应（DDR）的确定造血衰竭的进展 TERC的水平。特别是，我们将重点介绍p53途径在造血障碍中的作用端粒短的细胞，在异常造血和miR-145和miR-145的作用中解释了异常的造血和miR-145的作用破译沉默DDR以恢复具有DC-的细胞中造血输出的遗传后果相关突变。在目标两个方面，如果调节含有突变的细胞中的RNA衰减，我们将破译在DKC1中，是恢复这些细胞中造血输出的可行策略。我们将调制terc 通过抑制特定RNA衰变途径的转录后处理和降解。最后，在AIM 3中我们将调查TERC在端粒维护以外的新颖，非规范的作用是否会影响具有降低TERC的突变的患者的造血输出和增强疾病表型水平，因为这些通常会遭受严重疾病表型的折磨。总的来说，提出的研究将使我们能够破译针对定向疗法的新型靶标患有端粒合成的患者。我们独特的蜂窝和分子工具，结合我们端粒酶，干细胞生物学和造血的专业知识使我们处于理想的位置在这个领域的影响。