Functional Genomic Dissection of Refractory Anemia

难治性贫血的功能基因组解析

• 批准号: 8486470
• 负责人: Benjamin Levine Ebert
• 金额: $ 39.1万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-15 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8486470
• 关键词: 5q31; 5q32; Acute leukemia; Address; Alleles; Anemia; Animal Model; Apoptosis; Applications Grants; Binding; Biogenesis; Biological; Biological Assay; Biology; Blood; Bone Marrow; CD34 gene; Candidate Disease Gene; Cell Cycle Arrest; Cell physiology; Cells; Characteristics; Childhood; Chromosome Arm; Chromosome Deletion; Chromosomes; Chromosomes, Human, Pair 5; Code; Congenital Disorders; Defect; Development; Diamond-Blackfan anemia; Disadvantaged; Disease; Dissection; Dysmyelopoietic Syndromes; Epigenetic Process; Erythroid; Erythroid Progenitor Cells; Erythropoiesis; Failure; Funding; Gene Expression Profiling; Gene Targeting; Genes; Genetic; Genetic Engineering; Genetic Translation; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Human; In Vitro; Individual; Ineffective Hematopoiesis; Lead; Lesion; Link; MDM2 gene; Macrocytic Anemia; Malignant Neoplasms; Megakaryocytes; Messenger RNA; Methodology; MicroRNAs; Micromegakaryocyte; Modeling; Molecular; Molecular Abnormality; Mus; Oncogenes; Pancytopenia; Pathogenesis; Pathway interactions; Patients; Peripheral; Phenotype; Polyribosomes; Process; Production; Proteins; Protocols documentation; RNA; RNA Interference; RPS19 gene; Reading; Refractory anemias; Regulation; Relative (related person); Ribosomal Proteins; Ribosomes; Role; Stem cells; Study models; Sucrose; Syndrome; System; Technology; Testing; Time; Translating; Translations; Tumor Suppressor Proteins; Undifferentiated; United States; Validation; Zebrafish; base; chromosome 5q loss; chromosome 7q loss; clinical phenotype; functional genomics; genetic manipulation; in vivo; insight; interstitial; leukemia; novel; novel therapeutics; prevent; protein activation; public health relevance; rRNA Precursor; research study; screening; stem; thrombocytosis

DESCRIPTION (provided by applicant): Myelodysplastic syndrome (MDS) is characterized by ineffective hematopoiesis, most commonly of the erythroid lineage, resulting in a phenotype termed refractory anemia. In the 5q- syndrome, a subtype of MDS, a single genetic lesion, a heterozygous, interstitial deletion of Chromosome 5q, causes a highly reproducible clinical phenotype, though the molecular basis of this phenotype was previously unknown. In the previous funding period, we identified one protein-coding gene, RPS14, and one miRNA, miR-145, that contribute to abnormal hematopoiesis in the 5q- syndrome. The finding that RPS14 haploinsufficiency causes a block in erythropoiesis, the dominant phenotype of the 5q- syndrome, established a previously unrecognized link between the 5q- syndrome and Diamond Blackfan Anemia, a congenital disorder with a similar phenotype that is also caused by genetic inactivation of one allele of genes encoding ribosomal proteins. We found that haploinsufficiency for miR-145 causes increased expression of a key target gene, FLI-1, leading to increased megakaryocyte production and the characteristic hypolobated micromegakaryocytes found in this syndrome. In this renewal application, we aim to understand the molecular basis for the effects of RPS14 haploinsufficiency and combined haploinsufficiency for RPS14 and miR-145, and to examine the effects of these lesions on hematopoietic stem cells. In addition, having established an approach to the identification of key MDS genes within chromosomal deletions, we will apply our methodology to identify a key gene within the 7q deletion, another common genetic lesion in MDS. In Aim 1, we will investigate the mechanism whereby ribosomal haploinsufficiency leads to impaired erythropoiesis. Current evidence supports two non-exclusive hypotheses. The first possibility is that selective activation of p53 in the erythroid lineage causes cell cycle arrest and apoptosis, resulting in macrocytic anemia. Alternatively, or additionally, abnormal ribosome biogenesis could lead to dysregulated mRNA translation and altered production of specific proteins. We will examine both hypotheses in primary human bone marrow progenitor cells. In Aim 2, we will examine hematopoiesis in genetically engineered murine models with conditional haploinsufficiency of RPS14, miR-145, or the combination of RPS14 and miR-145. In particular, we will use these models to examine the effect of each lesion on hematopoietic stem cell function. In Aim 3, we will extend our RNA interference screening approach to identify additional genes that are critical for the pathogenesis of MDS. Having demonstrated the ability to use this approach to identify haploinsufficiency disease genes on Chromosome 5q, we will next focus this technology towards the identification of novel MDS genes on Chromosome 7q. In aggregate, these experiments will provide critical insight into the molecular basis of myelodysplastic syndrome.

描述（由申请人提供）：骨髓增生综合征（MDS）的特征是无效的造血症，最常见的是红斑谱系的，导致一种称为难治性贫血的表型。在5q-综合征中，MDS的亚型，单个遗传病变，5q染色体的杂合，间质缺失，导致高度重现的临床表型，尽管该表型的分子基础先前尚不清楚。在上一个资金期间，我们确定了一种蛋白质编码基因RPS14和一个miRNA，miRNA，它有助于5q-综合征中造血异常。 RPS14单倍弥平弥平液会导致5q-综合征的主要表型引起的障碍物，这一发现建立了先前未识别的5q-综合征与钻石黑芬贫血之间的连接，一种具有类似表型的先天性表型，这也是由类似的表型引起的，也是由基因构成基因的遗传灭菌引起的。我们发现，miR-145的单倍不足会导致关键靶基因FLI-1的表达增加，从而导致巨核细胞的产生增加和该综合征中发现的特征性降劳微瘤细胞。在这种续订应用中，我们旨在了解RPS14单倍体不足和RPS14和miR-145的综合单倍症的影响的分子基础，并检查这些病变对造血干细胞的影响。此外，在建立了鉴定染色体缺失中关键MDS基因的方法之后，我们将应用我们的方法来识别7Q缺失中的关键基因，这是MDS中另一种常见的遗传病变。在AIM 1中，我们将研究核糖体单倍体不足导致红细胞生成的机制。当前的证据支持两个非排他性假设。第一个可能性是，在红系谱系中p53的选择性激活会导致细胞周期停滞和凋亡，从而导致大细胞性贫血。或者，异常的核糖体生物发生可能导致mRNA翻译失调并改变了特定蛋白质的产生。我们将检查原代人骨髓祖细胞中的两个假设。在AIM 2中，我们将检查具有条件单倍不足的RPS14，miR-145或RPS14和miR-145的基因工程鼠模型中的造血菌素。特别是，我们将使用这些模型检查每个病变对造血干细胞功能的影响。在AIM 3中，我们将扩展RNA干扰筛选方法，以鉴定对MDS发病机理至关重要的其他基因。在证明了使用这种方法在5q染色体上鉴定单倍体疾病疾病基因的能力，我们接下来将把这项技术集中在鉴定新型MDS基因上的7q染色体上。总体而言，这些实验将为骨髓增生综合征的分子基础提供重要的见解。