Mechanisms of a GATA Factor-Dependent Hematopoietic Signaling Pathway

摘要 删除E-box-GATA内含子顺式元件(+9.5)可降低GATA-2的表达并使其失活 破坏GATA-2依赖基因的小鼠胚胎造血干细胞(HSC)生成器 网络。这个遗传网络的许多基本组成部分还没有被定义。去发现 将+9.5序列/分子性质与数千个+9.5-Like序列/分子性质进行比较 全基因组的顺式元件。这种创新的训练方法产生了一个+9.5类位点的全基因组文库， 含有GATA-2靶基因，可能通知独特的模式控制造血干细胞和 祖细胞(HSPC)。我发现GATA-2调节未研究过的不育α基序的表达 结构域14(Samd14)基因，Samd14可增加胎肝HSPC水平，并促进SCF/c-Kit信号转导。 初步数据表明，+9.5样Samd14-Enhancer介导Samd14的表达，是 应激红细胞生成，并使应激依赖的Samd14在脾中上调。这些目标将 严格建立Samd14促进SCF/c-Kit信号传导的机制(目标1)，Samd14的功能 GATA-2-调节Samd14-Enh在创新的小鼠模型(Aim 2)中，并定义了一组应激- 依赖基因及其互联性(目标3)。 作为一名K奖获得者，我将遵循详细的培训计划，并进行具体的研究和职业发展 在颁奖过程中要达到的里程碑。这些目标旨在扩大和区分我的 来自我导师的研究专业知识，我们已经同意我可以使用Samd14工作作为基础 发展我的独立研究项目。提案中概述的职业发展活动将 提供写作/演示技能、实验室管理和资质方面的培训。我组织了一次 指导委员会由久负盛名的成功科学家组成，并有相关的研究计划 对我的工作，他们致力于我的成功，并将在促进我的职业生涯过渡方面发挥积极作用。一个 该计划的主要内容包括直接培训骨髓移植技术以研究应激 造血术。威斯康星大学麦迪逊分校是一家顶级研究机构，拥有由美国国立卫生研究院赞助的优秀培训 项目、核心设施和研究实验室，这使它成为我理想的培训环境 进一步发展成为一名独立的科学家。我得到了威斯康星大学麦迪逊分校、The Cell和 再生生物系和Bresnick博士追求这些职业目标并竞争教员 在这里或其他研究密集型学术机构任职。研究目标、职业发展 培训计划、与指导委员会和合作者的会议、教育机会和培训 环境将确保发展的坚实基础，以启动富有成效的学术生涯。

Abstract Deleting an E-box-GATA intronic cis-element (+9.5) reduces GATA-2 expression and inactivates the hematopoietic stem cell (HSC) generator in the mouse embryo by disrupting GATA-2-dependent genetic networks. Many of the essential constituents of this genetic network have not been defined. To discover components of this network, the +9.5 sequence/molecular properties were compared to thousands of +9.5-like cis-elements genome-wide. This innovative training method yielded a genome-wide library of +9.5-like sites, containing GATA-2 target genes that may inform unique modes of controlling hematopoietic stem and progenitor cells (HSPCs). I discovered that GATA-2 regulates expression of the unstudied Sterile Alpha Motif Domain 14 (Samd14) gene, and Samd14 increased fetal liver HSPC levels and promoted SCF/c-Kit signaling. Initial data demonstrates that the +9.5-like Samd14-Enhancer mediates Samd14 expression, is required for stress erythropoiesis, and confers stress-dependent Samd14 upregulation in the spleen. These aims will rigorously establish the mechanism by which Samd14 promotes SCF/c-Kit signaling (Aim 1), the function of the GATA-2-regulated Samd14-Enh in an innovative mouse model (Aim 2), and define a cohort of stress- dependent genes and their interconnectivity (Aim 3). As a K award recipient, I will follow a detailed training plan with specific research and career development milestones to be achieved over the course of the award. The Aims are designed to expand and distinguish my research expertise from my mentor, and we have agreed that I can use the Samd14 work as a foundation for developing my independent research program. The career development activities outlined in the proposal will provide training in writing/presentation skills, laboratory management, and grantsmanship. I have organized a mentoring committee comprised of well-established and successful scientists, with research programs relevant to my work, who are committed to my success and will play an active role in facilitating my career transition. A key element of this plan entails direct training in bone marrow transplantation techniques to study stress hematopoiesis. UW-Madison is a top-tier research institution with outstanding NIH-sponsored training programs, core facilities, and research laboratories, which makes it an ideal training environment for me to further develop as an independent scientist. I have strong support from UW-Madison, the Cell and Regenerative Biology Department and Dr. Bresnick to pursue these career goals and compete for a faculty position here or at another research-intensive academic institution. The research Aims, career development training plan, meetings with mentoring committee and collaborators, educational opportunities, and training environment will ensure the development of a solid foundation to launch a productive academic career.