Regulation of blast cell quiescence by Pten and Tor

Pten 和 Tor 对母细胞静止的调节

• 批准号: 10335149
• 负责人: Julia Sarah Wittes
• 金额: $ 6.98万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10335149
• 关键词: Adult; Aging; Animals; Biological; Blast Cell; Brain; Caenorhabditis elegans; Cancer Biology; Cell Cycle; Cell Differentiation process; Cell Maintenance; Cell division; Cells; Complex; Development; Disease; Endocrine; Environment; Essential Genes; Functional disorder; Genes; Genetic; Genetic Screening; Genetic Techniques; Goals; Gonadal structure; Growth and Development function; Homeostasis; Homologous Gene; Human; Immunologic Surveillance; Insulin; Laboratories; Larva; Lipids; Maintenance; Malignant Neoplasms; Mammalian Cell; Modeling; Molecular; Mutate; Nematoda; Organism; Orthologous Gene; PTEN gene; Pathway interactions; Phenocopy; Phosphoric Monoester Hydrolases; Phosphotransferases; Protein phosphatase; Proteins; RNA interference screen; Regulation; Research; Research Personnel; Resistance; Signal Pathway; Signal Transduction; Sirolimus; Stress; Structure; System; Testing; Therapeutic; Time; Tissues; Training; Tumor Suppressor Proteins; Universities; adult stem cell; base; cancer stem cell; cancer therapy; cancer type; chemotherapy; design; experience; experimental study; feeding; fluorescence imaging; genetic analysis; germline stem cells; growth promoting activity; improved; in vivo; insight; insulin signaling; multipotent cell; mutant; nerve stem cell; protein complex; repaired; scaffold; stem; stem cell biology; stem cell function; stem cells; therapy development; tool; transcription factor; tumor

Project Summary/Abstract Quiescence is a defining feature of stem cells, which allows them to persist over time without losing developmental potential. Stem cells and other multipotent cells alternate between dormant quiescent periods and active periods of cell division and differentiation. However, our understanding of how these transitions are regulated is incomplete. The long-term objective of this project is to investigate how transitions between quiescence and differentiation are regulated by the genes Phosphatase and tensin homolog (Pten) and Target of Rapamycin (Tor), using the nematode worm C. elegans. In C. elegans, during a special larval stage called dauer, multipotent blast cells in the gonad are directed by signals from the environment to remain quiescent. The loss or maintenance of quiescence in these gonad blasts can easily be detected using fluorescent markers. Our laboratory recently showed that the C. elegans ortholog of Pten (DAF-18) regulates blast cell quiescence in the gonad of dauer larvae. The tumor suppressor Pten is an essential gene for mammalian development, and loss of Pten can cause many different types of cancer. Pten also regulates quiescence in adult mammalian stem cells and cancer stem cells, but we lack an understanding of how Pten does this. Quiescence is usually very difficult to study in vivo, and our gonadogenesis model provides a unique opportunity to study it in a living, intact organism, with unparalleled tools for genetic analysis. The goal of this research is to understand how, mechanistically, Pten promotes stem cell quiescence, using our gonadogenesis model. Pten/DAF-18 has multiple molecular activities: it is a protein phosphatase, a lipid phosphatase, and can also act as a ‘scaffold’ to assemble protein complexes. In Aim 1, structure/function studies will be used to assess which of these activities is required for DAF-18’s regulation of blast cell quiescence in our system. Additionally, to further understand DAF-18’s biological activity in regulating quiescence, the subcellular localization of endogenous DAF-18 protein in the gonad will be characterized. Aim 2 will use tissue-specific depletion experiments to test this hypothesis, based on preliminary experiments, that DAF-18 may promote cellular quiescence by opposing the activity of the growth-promoting Tor kinase pathway. In Aim 3, powerful genetic screens will be used to identify new genes that may act in a genetic pathway with DAF-18 to regulate quiescence. The experiments in this proposal will incorporate fluorescent imaging in living organisms and cutting-edge genetic techniques. This training is to be conducted at Columbia University over the course of two years, under the guidance of Dr. Iva Greenwald, a leading C. elegans geneticist with an excellent record of training successful researchers.

项目摘要/摘要 静止是干细胞的定义特征，它使它们能够随着时间的推移而持续而不会失去 发展潜力。干细胞和其他多能细胞在休眠静止期之间替代 和细胞分裂和分化的积极时期。但是，我们对这些过渡的理解 监管是不完整的。该项目的长期目标是调查如何 静态和分化受基因磷酸酶和tensin同源物（PTEN）和靶标的调节 雷帕霉素（Tor），使用线虫蠕虫秀丽隐杆线虫。在秀丽隐杆线虫中，在一个特殊的幼虫阶段称为 性腺中的多能爆炸细胞由环境的信号指向以保持静止。 可以使用荧光很容易检测到这些性腺爆炸中静止的损失或维持 标记。 我们的实验室最近表明，秀丽隐杆线虫的PTEN（DAF-18）正向手调节爆炸细胞的静止 在道尔幼虫的性腺中。肿瘤抑制剂PTEN是哺乳动物发育的重要基因， PTEN的丧失会导致许多不同类型的癌症。 PTEN还调节成人哺乳动物的静止 干细胞和癌症干细胞，但我们对PTEN的执行方式缺乏了解。静止通常是 非常困难的体内研究，我们的性腺生成模型为研究它提供了独特的机会， 完整的生物，具有无与伦比的遗传分析工具。这项研究的目的是了解如何 从机械上讲，PTEN使用我们的性腺生成模型促进了干细胞静止。 PTEN/DAF-18具有多种分子活性：它是一种蛋白质磷酸酶，一种脂质磷酸酶，也可以 充当组装蛋白质复合物的“支架”。在AIM 1中，结构/功能研究将用于评估 DAF-18在我们系统中对爆炸细胞静止的调节需要哪些活动。此外， 为了进一步了解DAF-18在控制静止方面的生物学活性， 性腺中的内源性DAF-18蛋白将被表征。 AIM 2将使用组织特异性部署 基于初步实验测试该假设的实验，DAF-18可能会促进细胞 通过反对促进生长的TOR激酶途径的活性来静止。在AIM 3中，强大的遗传 屏幕将用于识别可能在DAF-18的遗传途径中起作用的新基因以调节 静止。该提案中的实验将在生物体中结合荧光成像， 尖端的遗传技术。这项培训将在两个过程中在哥伦比亚大学进行 多年来，在伊瓦·格林瓦尔德（Iva Greenwald）博士的指导下，伊瓦·格林瓦尔德（Iva Greenwald 培训成功的研究人员。