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）：这项研究的长期目标是揭示基因表达如何在转录后级别调节。信使RNA（mRNA）翻译和降解的控制是重要的生物学过程，包括发育，生育和神经功能。拟议的工作着重于原型mRNA调节剂Pumilio（PUM）。 PUM是真核RNA结合蛋白的PUF家族的成员。 PUM结合了一组广泛的使者RNA，其中包括编码细胞周期蛋白和关键发育形态的那些。与mRNA结合后，PUM抑制编码蛋白的表达。拟议的研究旨在发现PUM抑制的机制，并确定如何调节PUM活动。我们开发了新的测定方法，以测量果蝇细胞中的脓液活性，并发现了有效抑制蛋白质表达的多个新域。 PUM抑制域的抑制活性不依赖于先前鉴定的辅助因子，表明新型的抑制机制。在第一个目标中，我们衡量抑制域对翻译和mRNA降解的影响。我们试图发现其活动所需的辅助因子。我们发现了一个自动调节开关，该开关控制了PUM活性，可以通过磷酸化调节。在第二个目标中，我们使用功能和蛋白质相互作用测定法剖定了这种自动调节开关的机制。在第三个目标中，我们探讨了Zn手指蛋白纳米如何通过PUM调节抑制作用。纳米与PUM相互作用并增强抑制作用。我们建议纳米增强PUM与mRNA的相互作用，并实际上改变了由PUM抑制的mRNA的收集。这项研究将确定PUM抑制的新型机制，并阐明如何根据蛋白质修饰和与纳米的结合作用来调节该活性。预计所产生的发现将广泛增强我们对转录后控制的理解，并专门提高对发育，生育能力，干细胞增殖和神经系统中基因调节的了解。