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.

项目概要/摘要 静止是干细胞的一个决定性特征，这使得它们能够随着时间的推移而持续存在而不会丢失 干细胞和其他多能细胞在休眠静止期之间交替。 然而，我们对这些转变是如何进行的理解。 该项目的长期目标是研究两者之间的过渡。 静止和分化由磷酸酶和张力蛋白同源物 (Pten) 和靶标基因调节 雷帕霉素 (Tor)，使用线虫秀丽隐杆线虫，在称为线虫的特殊幼虫阶段。 dauer，性腺中的多能母细胞在环境信号的引导下保持静止状态。 使用荧光可以轻松检测到这些性腺母细胞的静止状态的丧失或维持 标记。 我们的实验室最近表明，Pten (DAF-18) 的线虫直系同源物可调节母细胞静止 在 dauer 幼虫的性腺中，肿瘤抑制基因 Pten 是哺乳动物发育的必需基因， Pten 的缺失可能导致许多不同类型的癌症，Pten 还调节成年哺乳动物的静止状态。 干细胞和癌症干细胞，但我们对 Pten 通常如何做到这一点缺乏了解。 在体内研究非常困难，而我们的性腺发生模型提供了在活体中研究它的独特机会， 完整的有机体，具有无与伦比的遗传分析工具，这项研究的目标是了解如何， 从机制上讲，Pten 利用我们的性腺发生模型促进干细胞静止。 Pten/DAF-18具有多种分子活性：它是一种蛋白磷酸酶、一种脂质磷酸酶，还可以 充当组装蛋白质复合物的“支架” 在目标 1 中，结构/功能研究将用于评估。 其中哪些活性是 DAF-18 调节我们系统中母细胞静止所必需的。 为了进一步了解 DAF-18 在调节静止方面的生物活性， 目标 2 将使用组织特异性去除来表征性腺中的内源性 DAF-18 蛋白。 实验验证这一假设，基于初步实验，DAF-18 可能促进细胞 在目标 3 中，通过对抗生长促进 Tor 激酶途径的活性来实现静止。 筛选将用于识别可能与 DAF-18 一起在遗传途径中发挥作用的新基因，以调节 该提案中的实验将在生物体中结合荧光成像和 该培训将在哥伦比亚大学进行，为期两年。 多年来，在 Iva Greenwald 博士的指导下，Iva Greenwald 博士是一位领先的线虫遗传学家，在 培训成功的研究人员。