Determining piwi function using in vivo imaging of stem cells in Hydra.

使用 Hydra 干细胞的体内成像确定 piwi 功能。

基本信息

批准号：
8206625
负责人：
Celina Juliano
金额：
$ 5.22万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-01-04 至 2013-01-03
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Stem cells are capable of both self-renewal and producing numerous differentiated cells. The Piwi subfamily of Argonaute proteins (herein called Piwi proteins) is required for stem cell maintenance in diverse organisms, yet its molecular function is not well understood. Piwi proteins associate with a distinct class of small RNAs, piRNAs, and modulate gene expression at both epigenetic and post-transcriptional levels. This proposal will take advantage of Hydra as a model system to further understand piwi functions in stem cells that are conserved across phylogeny. The Hydra, a basal metazoan, has a well-characterized stem cell lineage, the 1-cell lineage, in which two piwi homologs are specifically expressed. Transgenic Hydra that express GFP in the 1-cell lineage allow for in vivo observation of stem cells, thus, for the first time, the effect of piwi perturbation on stem cell behavior will be observed in the context of a living organism. In addition, because Hydra propagate clonally it is easy to obtain large, genetically pure populations for biochemical studies. Thus, the molecular function of piwi in stem cells will be investigated by characterizing PlWl-containing protein complexes and identifying targets of piwi-directed gene regulation. Finally, Hydra has a large population of putative piRNAs of which the identity and function will be explored. Importantly, Cnidarian gene sets, such as Hydra, exhibit the same complexity found in vertebrate genomes. Thus, functional discoveries made in these relatively simple metazoans will be applied to more complex organisms. These studies will advance our understanding of the conserved molecular functions of piwi in stem cells, thus contributing to the basic understanding of stem cell molecular circuitry required to manipulate stem cells for use in therapy. PUBLIC HEALTH RELEVANCE: Stem cells both self-renew and give rise to differentiated cell types of the body and therefore have great therapeutic potential in replacing lost or damaged tissues. The piwi genes are required for stem cell self-renewal in diverse organisms; they encode proteins that associate with small RNAs to modulate gene expression at many levels. Hydra, with its accessibility for manipulating stem cells in the context of a live organism, provides an effective model to explore molecular mechanisms mediated by Piwi proteins in regulating stem cell self-renewal and differentiation.

描述（由申请人提供）：干细胞能够自我更新并产生许多分化的细胞。 Argonaute蛋白的PIWI亚家族（以下称为PIWI蛋白）是多种生物中干细胞维持所必需的，但其分子功能尚不清楚。 PIWI蛋白与表观遗传和转录后水平的一类不同类别的RNA，PIRNA和调节基因表达相关。该建议将利用Hydra作为模型系统，以进一步了解在系统发育中保守的干细胞中PIWI功能。九头蛇是一种基底后生动物，具有一个特征良好的干细胞谱系，即1细胞谱系，其中有两个Piwi同源物被专门表达。在1细胞谱系中表达GFP的转基因HydrA可以在体内观察干细胞，因此，在生物生物体的背景下，将观察到Piwi扰动对干细胞行为的影响。此外，由于九头蛇在克隆上繁殖，因此很容易获得大型的，遗传上的纯种种群进行生化研究。因此，将通过表征含有PLWL的蛋白质复合物并鉴定PIWI定向基因调节靶标的PIWI在干细胞中的分子功能。最后，Hydra有大量的推定PIRNA，将探索其身份和功能。重要的是，诸如HYDRA之类的cNidarian基因集具有在脊椎动物基因组中发现的复杂性。因此，这些相对简单的后生动物中的功能性发现将应用于更复杂的生物体。这些研究将促进我们对干细胞中PIWI保守的分子功能的理解，从而有助于对操纵干细胞用于治疗所需的干细胞分子电路的基本理解。公共卫生相关性：干细胞自我更新并引起人体的分化细胞类型，因此在替换损失或受损的组织方面具有巨大的治疗潜力。 PIWI基因在多种生物体中需要干细胞自我更新所必需。它们编码与小RNA相关的蛋白质以在许多级别调节基因表达。 Hydra具有在活生物体的背景下操纵干细胞的可访问性，它提供了一个有效的模型，以探索PIWI蛋白在调节干细胞自我更新和分化方面介导的分子机制。