Mechanisms governing stem cell coordination by the niche

通过生态位控制干细胞协调的机制

• 批准号: 10389225
• 负责人: Kari Lenhart
• 金额: $ 10万
• 依托单位: DREXEL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10389225
• 关键词: 3-Dimensional; Administrative Supplement; Biological Models; Cell Communication; Cell Lineage; Cell physiology; Complex; Cytokinesis; Data; Daughter; Drosophila genus; Ensure; Environment; Excision; F-Actin; Funding; Genetic; Germ Cells; Goals; Hair follicle structure; Hematopoietic; Homeostasis; Hour; Image; Laser Scanning Confocal Microscopy; Lasers; Lateral; Maintenance; Microscope; Modification; Molecular; Molecular Genetics; Outcome; Output; Ovary; Population; Production; Productivity; Resolution; Scanning; Signal Transduction; Source; Structure; System; Testis; Time; Tissues; Tumor Stem Cells; Work; adult stem cell; cell type; daughter cell; detector; genetic manipulation; germline stem cells; imaging capabilities; improved; insight; instrumentation; parent grant; prevent; programs; quantum; reconstruction; spatiotemporal; sperm cell; stem cell population; stem cell proliferation; stem cells

Project Summary / Abstract Coordination between adult stem cells is essential to maintain tissue homeostasis and prevent tumorous overgrowth. Many structures, including the hair follicle, hematopoietic network and developing ovary require tight control over stem cell proliferation and coordination of daughter cell production from distinct stem cell lineages. In most cases, the molecular mechanisms orchestrating this coordination are largely unknown. Leveraging the power of Drosophila genetics and establishing a system for longitudinal (20+ hours) live imaging of stem cells within an endogenous niche we have begun to reveal the mechanisms controlling stem cell coordination in the testis. Somatic stem cells and germline stem cells (GSCs) of the testis must generate daughters in a precise 2:1 ratio for germ cells to effectively differentiate into sperm. Our live imaging has revealed a modified cytokinesis program in GSCs as the mechanism to coordinate release of one GSC daughter only after it correctly associates with two daughters of the somatic stem cell lineage. This modified cytokinesis program is controlled at two stages—a pause regulated by Jak/STAT signaling from the niche and a trigger for completion of cytokinesis derived from the somatic stem cells. Both control points must be properly executed or stem cell cytokinesis fails, stem cell tumors form and germ cells fail to differentiate. While we have identified the source of both the pause and trigger, the mechanisms by which these signals control GSC cytokinesis remain unknown. In the parent grant, we propose to interrogate the specific mechanisms by which niche signals and somatic stem cells combine to regulate GSC cytokinesis using molecular genetics and extended live imaging. Here, we request an administrative supplement for the acquisition of fast and super-resolution laser scanning confocal microscopy, which will improve the imaging capabilities, quality and data output (productivity) of our R01-funded projects. The new instrumentation will enable imaging of fixed and live testes faster and with less photodamage as well as unprecedented resolution (100 nm lateral, 200 nm axial). This enhanced resolution will aid in identification of altered F-actin structure at the intercellular bridge between GSC-daughter pairs investigated in Aim1 of the parent grant. In our studies of temporal dynamics in abscission machinery localization to the GSC-daughter intercellular bridge (Aim2 of parent grant), the new instrumentation will enable tracking of ESCRT machinery with unprecedented spatiotemporal resolution in 2D and 3D using detectors with superior quantum efficiency. Through the use of advanced modulations such as dynamic enhancement and adaptive image quality determination and reconstruction, we will be able to acquire high-quality time-lapse data with improved resolution. This is in conjunction with the ability to acquire images from multiple (up to three) wavelengths simultaneously—an essential feature for our work dissecting the complex interactions between somatic and germline stem cells through live imaging of both populations (Aim3 of parent grant). Outcomes will shed valuable insight into niche-regulated cytokinesis modifications in stem cells. In addition, this work will provide the first real-time, high-resolution analysis of stem cell interactions within an endogenous niche that are essential for maintenance of tissue homeostasis.

项目摘要/摘要 成体干细胞之间的协调对于维持组织动态平衡和预防 肿瘤性过度生长。许多结构，包括毛囊、造血网和 卵巢发育需要严格控制干细胞增殖和子代的协调 从不同的干细胞谱系产生细胞。在大多数情况下，分子机制 协调这种协调在很大程度上是未知的。利用果蝇的力量 遗传学和建立干细胞纵向(20多小时)活体成像系统 一个内源性的利基我们已经开始揭示控制干细胞的机制 睾丸的协调性。睾丸的体细胞和生殖系干细胞(GSC)必须 以精确的2：1比例产生后代，使生殖细胞有效地分化为精子。 我们的实时成像揭示了GSCs中一种修改的胞质分裂程序作为 协调释放一个GSC子项，前提是该子项正确关联到 体细胞干细胞谱系。这个修改后的胞质分裂程序被控制在两个阶段--a 暂停由来自缝隙的JAK/STAT信号调节，并触发完成 来自体细胞干细胞的胞质分裂。两个控制点都必须正确执行 或者干细胞胞质分裂失败，干细胞肿瘤形成，生殖细胞无法分化。当我们 已经确定了暂停和触发的来源，以及通过这些机制 控制GSC胞质分裂的信号仍不清楚。在家长拨款中，我们建议审问 利基信号和躯体干细胞联合调控的具体机制 使用分子遗传学和扩展的活体成像进行GSC胞质分裂。在这里，我们请求一个 获取快速和超分辨率激光扫描的行政补充 共焦显微镜，将提高成像能力、质量和数据输出 (生产力)我们由R01资助的项目。新的仪器将使固定的成像 和现场测试更快，光损伤更小，以及前所未有的分辨率(100 nm 横向，200 nm轴向)。这种增强的分辨率将有助于识别改变的F-肌动蛋白 在Aim1中研究的GSC-子代对之间的细胞间桥结构 家长助学金。在我们对剥离机械中的时间动力学的研究中，定位于 GSC-子代细胞间桥(父母GRANT的AIM2)，新的仪器将启用 在2D和3D中以前所未有的时空分辨率跟踪ESCRT机器 具有卓越量子效率的探测器。通过使用高级调制，例如 动态增强和自适应图像质量的确定和重建，我们将 能够以更高的分辨率获取高质量的延时数据。这是与 能够同时从多个(最多三个)波长获取图像- 我们解剖体细胞和生殖系之间复杂相互作用的基本特征 干细胞通过两个群体的活体成像(父母赠款的Aim3)。结果将会消失 对干细胞中利基调节的胞质分裂修饰的有价值的见解。此外，这一点 这项工作将提供对干细胞相互作用的第一次实时、高分辨率分析 维持组织内环境平衡所必需的内源性生态位。