Zebrafish Genetics to Identify Genes Involved in Hematopoiesis

斑马鱼遗传学鉴定参与造血的基因

基本信息

批准号：
8531909
负责人：
DHVANIT INDRAVADAN SHAH
金额：
$ 15.86万
依托单位：
BRIGHAM AND WOMEN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-15 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8531909
关键词：
ATP phosphohydrolase Adult Affect Alleles Aminolevulinic Acid Anemia Animal Model Award Biochemical Biogenesis Biological Assay Biology Blood Blood Vessels Candidate Disease Gene Cell Count Cell Differentiation process Cells Child Chromosome Mapping Communities Complex Congenital Abnormality Data Defect Development Developmental Biology Dianisidine Diploidy Embryo Environment Erythrocytes Erythroid Erythroid Cells Erythropoiesis Erythropoietic Protoporphyria Exhibits Face Finding of Mean Corpuscular Hemoglobin Foundations Functional disorder Future Genes Genetic Genetic Markers Genetic Screening Genetic Transcription Globin Goals Haploidy Hematocrit procedure Hematological Disease Hematopoiesis Hematopoietic Heme Hemochromatosis Hemoglobin Human Human Genetics Human Genome Hypochromic anemia Iron Iron Metabolism Disorders Knowledge Lesion Link Medical Mentored Research Scientist Development Award Mentors Mesoderm Cell Mitochondria Modeling Molecular Mutagenesis Myelogenous National Institute of Diabetes and Digestive and Kidney Diseases Nitrosourea Compounds Organ Outcome Pathway interactions Poisson Distribution Randomized Regulation Research Personnel Role Scientist Signal Transduction Staining method Stains Stem Cell Development Stem cells Synteny Time Variant World Health Organization Zebrafish base effective therapy genetic analysis hematopoietic tissue improved insight iron metabolism loss of function mRNA Expression mutant novel positional cloning progenitor public health relevance self-renewal stem success tool

项目摘要

DESCRIPTION (provided by applicant): The goal of this project is to offer novel insights into heme synthesis and hematopoietic pathways that can improve treatment of congenital and acquired anemia. Despite decades of efforts to understand genetic blood disorders, every year 400,000 newly born babies worldwide face dreadful complications in the absence of effective therapies. Thus, better understanding and knowledge of heme synthesis, iron metabolism, and hematopoietic stem/progenitor revcell (HSPC) biogenesis is required. The zebrafish, with its close synteny to the human genome and its conserved molecular pathways regulating the development of hematopoietic tissues, offers a powerful tool to understand human blood disorders. Based on our Poisson distribution analyses, we hypothesized that the genetic screens for anemic mutants are not at saturation to study hematologic diseases. We employed de novo ethyl nitrosourea (ENU) based genetic screens and identified five unique zebrafish anemic mutants, including sauvignon blanc (sav) and pinotage (pnt). pnt exhibits hypochromic anemia. pnt has a defect in hemoglobinisation while preserving red cell numbers. In contrast, sav shows anemia due to a lack of erythroid cells. sav lacks expression markers for HSPC (scl/tal1), myeloerythroid progenitor (gata-1), erythroid ((-globin) and myeloid (mpo) cells. Expression of vascular markers such as, fli-1 is preserved in sav. These data suggest that the pathophysiology of sav is confined to hematopoietic cell development and not to mesodermal cells. The genetic mapping, positional cloning, loss-of-function, mRNA expression levels, and cRNA over-expression studies have revealed a novel functional gene for the pnt locus. We have identified closely linked (~0 cM) genetic markers flanking the unique sav locus. In order to further characterize the importance of the disrupted genes in pnt and sav mutants and their involvement in genetic blood disorders, we propose: 1) To analyze the biochemical function of the pnt gene in heme synthesis. The pathophysiology of the pnt gene in hypochromic anemia will be ascertained by analyzing mechanisms regulating transport and incorporation of iron into heme or [Fe-S] cluster synthesis using mammalian and other model organisms. 2) To identify the sav gene and its function in hematopoiesis. The positional cloning, loss-of-function, cRNA over-expression studies and mutational analyses will be performed to identify the sav gene. We will perform assays to derive cell intrinsic or extrinsic role of sav in the regulation of hematopoiesis, hematopoietic stem/progenitor cell development, stem cell self-renewal, and cell differentiation. In the long term, the successful outcomes of these projects will lay strong foundations to understand human genetic blood diseases and to discover new opportunities for improved therapies. The potential of zebrafish genetics, expert guidance of mentors and scientific advisors, and the environment of Harvard Medical Community will certainly provide the candidate a unique opportunity to evolve as a successful independent investigator.

描述（由申请人提供）：该项目的目的是提供有关血红素合成和造血途径的新见解，这些途径可以改善先天性和获得性贫血的治疗。尽管几十年来努力了解遗传疾病，但在没有有效疗法的情况下，每年有400,000名全世界的新生婴儿面临可怕的并发症。因此，需要更好地理解和了解血红素合成，铁代谢和造血干/祖细胞Revcell（HSPC）生物发生。斑马鱼与人类基因组及其对调节造血组织发展的保守分子途径的同步，提供了一种强大的工具来了解人类血液疾病。基于我们的泊松分布分析，我们假设贫血突变体的遗传筛选不饱和研究血液学疾病。我们采用了基于新乙基硝基（ENU）的遗传筛选，并确定了五个独特的斑马鱼贫血突变体，包括长相思（SAV）和Pinotage（PNT）。 PNT表现出低色素性贫血。 PNT在保留红细胞数量的同时，血红蛋白缺陷。相反，由于缺乏红系细胞，SAV显示出贫血。 sav lacks expression markers for HSPC (scl/tal1), myeloerythroid progenitor (gata-1), erythroid ((-globin) and myeloid (mpo) cells. Expression of vascular markers such as, fli-1 is preserved in sav. These data suggest that the pathophysiology of sav is confined to hematopoietic cell development and not to mesodermal cells. The genetic mapping,位置克隆，功能丧失，mRNA表达水平和CRNA的过表达研究表明，我们的PNT基因座的新功能性基因已确定，我们已经识别出独特的SAV基因座的紧密联系（〜0 cm），以进一步表征其在PNT和SAVEN中的重要性。 PNT基因在血红素合成中的生化功能。低色素性贫血中PNT基因的病理生理将通过分析使用哺乳动物和其他模型生物来调节铁和[Fe-s]簇合成中的转运和掺入的转运和掺入的机制来确定。 2）确定SAV基因及其在造血中的功能。将进行位置克隆，功能丧失，CRNA过表达研究和突变分析，以识别SAV基因。我们将执行SAV在调节造血，造血干/祖细胞发育，干细胞自我更新和细胞分化中的固有或外在作用的测定。从长远来看，这些项目的成功结果将奠定强大的基础，以了解人类遗传疾病，并发现改善疗法的新机会。斑马鱼遗传学，导师和科学顾问的专家指导以及哈佛医学界的环境的潜力肯定会为候选人提供一个独特的机会，以发展为成功的独立研究者。