Transcriptional regulatory networks in spermatogonial stem cells

精原干细胞的转录调控网络

• 批准号: 8494137
• 负责人: Brian Peter Hermann
• 金额: $ 8.23万
• 依托单位: UNIVERSITY OF TEXAS SAN ANTONIO
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8494137
• 关键词: Address; Adult; Area; Binding; Bioinformatics; Biological; Biological Assay; Cell Culture Techniques; Cells; Commit; Complex; Cytokinesis; Data; Data Analyses; Development; Development Plans; Doctor of Philosophy; EMSA; Environment; Faculty; Fertility; Foundations; Funding; Future; Gene Combinations; Gene Expression; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Transcription; Germ Cells; Goals; Infertility; Instruction; K-Series Research Career Programs; Letters; Life; Luciferases; Male Infertility; Mentors; Mentorship; Methods; Mitosis; Molecular; Mus; Pathway interactions; Pattern; Phase; Plants; Positioning Attribute; Primates; Principal Investigator; Process; Production; Productivity; Program Evaluation; Property; Proteins; Regulation; Regulator Genes; Regulatory Element; Reporter; Reproductive Biology; Reproductive Physiology; Research; Research Institute; Research Personnel; Resources; Rodent; Role; Running; Science; Secure; Site; Spermatogenesis; Spermatogonia; Stem cell transplant; Stem cells; Structure; Subfamily lentivirinae; T-cell acute lymphocytic leukemia 1 protein; Testing; Testis; Tissues; Training; Transcriptional Regulation; Transfection; Translations; Transplantation; Undifferentiated; Universities; Walkers; Woman; Work; ZNF145 gene; abstracting; base; career; career development; cell behavior; chromatin immunoprecipitation; cohort; design; embryonic stem cell; experience; in vivo; innovation; insight; interest; male; medical schools; member; mouse model; network models; novel strategies; post-doctoral training; progenitor; programs; promoter; research study; self-renewal; sertoli cell; sperm cell; stem; stem cell biology; stem cell fate; transcription factor

Project Summary/Abstract Spermatogonial stem cells (SSCs) are at the foundation of mammalian spermatogenesis, maintaining sperm production throughout adult life. The molecular mechanisms that control the critical self-renewal vs. differentiation fate decision of SSCs are largely unknown. The hypothesis is that transcription factors primarily restricted to stem and progenitor spermatogonia form regulatory networks to execute gene expression programs important for SSC fate decisions and spermatogenesis. Three Specific Aims will test this hypothesis. Experiments in Specific Aim 1 will employ functional genetics to determine if specific transcription factors are essential for spermatogenesis using a knockdown strategy in ex vivo SSC cultures followed by transplantation to assess stem cell and spermatogenic activity. Specific Aim 2 will identify the target gene repertoire of SSC transcription factors by ChIP-Seq and reveal conserved and unique targets sets. Experiments in Specific Aim 3 will confirm mechanisms of target gene regulation by EMSA and transient transfection transcriptional analysis. These results will form the basis of gene regulatory network models incorporating complex interactions from multiple factors contributing to gene regulation and SSC potential. The proposed studies will be enhance the independent career trajectory of the PI (Brian Hermann) by establishing a new research focus investigating the fundamental regulation of SSCs in the rodent testis under the mentorship of Dr. Kyle Orwig. This is a new research direction for the PI who has established expertise in Sertoli cell gene regulation and primate spermatogenesis and stem cells. The intellectual and technical environment for research in reproductive biology and stem cells at the Magee-Womens Research Institute and University of Pittsburgh is outstanding, all resources required to complete the studies are available, and there is a strong reputation for promoting development of new investigators. This combination of a PI with outstanding potential for independence, an innovative mentor, and superior environment will facilitate all aspects of the proposed studies and advancement of PI's career towards independence. The long-term career goals of the PI are to establish an independent research program in a tenure-track faculty position, make significant scholarly contributions to the understanding of SSCs and spermatogenesis, and develop new approaches for treating male infertility. Three immediate objectives to promote the transition to independence are addressed by the career development award: 1) develop an independent research niche, 2) demonstrate outstanding productivity, and 3) secure additional independent funding. The career development plan involves conceptual mentoring in functional genetics, technical training relating to culture and transplantation of SSCs in the mouse model, instruction in bioinformatics data analysis, informal and structured mentoring in areas relevant to running a successful independent research program, and evaluation of career development progress by an external advisory board. Ultimately, these studies may provide insights about the mechanisms that initiate and maintain spermatogenesis, which has implications for treating male infertility. Investigating the biological properties of SSCs may also expand the understanding of how stem cells behave and contribute to the normal function of a variety of adult tissues. Moreover, additional nurturing of the candidate's career development will promote a competitive, independent research career to contribute substantively to the biomedical sciences.

项目摘要/摘要 精原干细胞(SSCs)是哺乳动物精子发生的基础， 在成年后的整个生命中产生精子。控制关键的自我更新的分子机制。 SSCs的分化命运决定在很大程度上是未知的。假设转录因子主要是 限制于干和祖细胞的精原细胞形成调控网络来执行基因表达 对SSC命运决定和精子发生很重要的程序。三个具体目标将检验这一假设。 在特定目标1的实验将使用功能遗传学来确定特定的转录因子 在体外SSC培养中使用敲除策略对精子发生至关重要，随后 移植以评估干细胞和生精活性。特定目标2将确定目标基因 ChIP-Seq的SSC转录因子谱，揭示保守和独特的靶点集。 特定目标3的实验将确认EMSA和瞬变对靶基因的调控机制 转导转录分析。这些结果将构成基因调控网络模型的基础 融合了多种因素的复杂相互作用，这些因素有助于基因调控和SSc潜力。 拟议的研究将通过以下方式增强PI(布莱恩·赫尔曼饰)的独立职业轨迹 建立一个新的研究热点，研究啮齿动物睾丸中SSCs的基本调控 凯尔·奥维格博士的指导。这是一个新的研究方向，对于已经建立了专业知识的 支持细胞基因调控与灵长类精子发生和干细胞。知识和技术 玛吉妇女研究所生殖生物学和干细胞研究的环境和 匹兹堡大学是杰出的，完成学业所需的所有资源都可用，而且在那里 有很强的口碑推动新的调查员的发展。PI与 突出的独立潜力、创新的导师和优越的环境将促进 拟开展的研究工作的方方面面，以及推动国际和平研究所走向独立的事业。 PI的长期职业目标是在终身教职轨道上建立一个独立的研究计划 在理解干细胞和精子发生方面做出了重大的学术贡献， 并开发治疗男性不育的新方法。促进过渡的三个近期目标 职业发展奖针对的是独立：1)发展独立的研究利基市场， 2)表现出出色的工作效率，3)获得额外的独立资金。职业生涯 发展计划包括功能遗传学的概念指导、与文化有关的技术培训 和在小鼠模型中移植SSCs，生物信息学数据分析指导，非正式和 在与成功运行独立研究计划和评估相关的领域进行结构化指导 由外部咨询委员会对职业发展进展情况进行评估。 归根结底，这些研究可能提供关于启动和维持 精子发生，这对治疗男性不育有意义。研究细菌的生物学特性 SSCS还可能扩大对干细胞如何行为的理解，并有助于 各种成人组织。此外，对应聘者职业发展的额外培养将促进 竞争激烈、独立的研究生涯，为生物医学科学做出实质性贡献。