Request to Transfer R01GM105707-03 Novel regulatory mechanisms of Drosophila Pumilio and Nanos

请求转让 R01GM105707-03 果蝇 Pumilio 和 Nanos 的新颖调控机制

基本信息

批准号：
9320859
负责人：
Aaron Charles Goldstrohm
金额：
$ 28.85万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-20 至 2018-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9320859
关键词：
Address Binding Binding Sites Biochemical Biological Biological Assay Biological Process Cancer Biology Cell Cycle Proteins Cell Proliferation Cells Collection Defect Development Drosophila genus EIF4EL3 gene Engineering Enzymes Failure Family Fertility Fingers Gene Expression Gene Expression Regulation Genes Genetic Genetic Transcription Goals Health Homeostasis Human Human Biology Infertility Knowledge Learning Locomotion Measures Messenger RNA Modeling Nervous System Physiology Nervous system structure Phenotype Phosphorylation Positioning Attribute Post-Transcriptional Regulation Post-Translational Protein Processing Process Proteins RNA Binding RNA Decay RNA Degradation RNA Recognition Motif RNA-Binding Proteins Recruitment Activity Regulation Repression Research Role Specificity Stem cells System Therapeutic Transcription Repressor/Corepressor Translations Work Zinc Fingers base cofactor combinatorial genetic regulatory protein improved insight member morphogens mutant nano nervous system disorder novel protein expression public health relevance response tool

项目摘要

DESCRIPTION (provided by applicant): The long term goal of this research is to reveal how gene expression is regulated at the post-transcriptional level. Control of messenger RNA (mRNA) translation and degradation underlies important biological processes including development, fertility and neurological functions. The proposed work focuses on the archetypal mRNA regulator, Pumilio (Pum). Pum is a member of the PUF family of eukaryotic RNA binding proteins. Pum binds an extensive group of messenger RNAs including those that encode cell cycle proteins and key developmental morphogens. Upon binding to an mRNA, Pum represses expression of the encoded protein. The proposed research seeks to discover the mechanism of Pum repression and determine how Pum activity is modulated. We developed novel assays to measure Pum activity in Drosophila cells and discovered multiple new domains that potently repress protein expression. The repressive activity of the Pum repression domains do not rely on previously identified cofactors, indicating novel repression mechanism(s). In the first aim, we measure the impact of the repression domains on translation and mRNA degradation. We seek to discover cofactors necessary for their activity. We discovered an autoregulatory switch that controls Pum activity and may be modulated by phosphorylation. In the second aim, we dissect the mechanism of this autoregulatory switch using functional and protein interaction assays. In the third aim, we explore how the Zn finger protein Nanos regulates repression by Pum. Nanos interacts with Pum and enhances repression. We propose that Nanos enhances interaction of Pum with mRNA and, in effect, changes the collection of mRNAs repressed by Pum. This research will determine novel mechanisms of Pum repression and illuminate how that activity is regulated in response to protein modifications and combinatorial action with Nanos. The resulting discoveries are expected to broadly enhance our understanding of post-transcriptional control and specifically improve knowledge of gene regulation in development, fertility, stem cell proliferation, and the nervous system.

描述（由申请人提供）：本研究的长期目标是揭示转录后水平上基因表达的调控方式。信使 RNA (mRNA) 翻译和降解的控制是发育、生育和神经功能等重要生物过程的基础。拟议的工作重点是原型 mRNA 调节因子 Pumilio (Pum)。 Pum 是真核 RNA 结合蛋白 PUF 家族的成员。 Pum 结合大量信使 RNA，包括编码细胞周期蛋白和关键发育成形素的信使 RNA。与 mRNA 结合后，Pum 会抑制所编码蛋白质的表达。拟议的研究旨在发现 Pum 抑制机制并确定 Pum 活性是如何调节的。我们开发了新的测定方法来测量果蝇细胞中的 Pum 活性，并发现了多个有效抑制蛋白质表达的新结构域。 Pum 抑制域的抑制活性不依赖于先前识别的辅助因子，表明新的抑制机制。第一个目标是测量抑制域对翻译和 mRNA 降解的影响。我们寻求发现其活性所需的辅助因子。我们发现了一种控制 Pum 活性并可能通过磷酸化进行调节的自动调节开关。在第二个目标中，我们使用功能和蛋白质相互作用分析来剖析这种自动调节开关的机制。第三个目标是探索 Zn 指蛋白 Nanos 如何调节 Pum 的抑制。 Nanos 与 Pum 相互作用并增强抑制作用。我们认为 Nanos 增强了 Pum 与 mRNA 的相互作用，并且实际上改变了 Pum 抑制的 mRNA 的集合。这项研究将确定 Pum 抑制的新机制，并阐明如何响应蛋白质修饰和与 Nanos 的组合作用来调节该活性。由此产生的发现预计将广泛增强我们对转录后控制的理解，并特别提高对发育、生育力、干细胞增殖和神经系统中基因调控的了解。