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RNA Regulation of Transcription Factor Activity

RNA对转录因子活性的调节

基本信息

批准号：
10316936
负责人：
DEBORAH S. WUTTKE
金额：
$ 49.85万
依托单位：
UNIVERSITY OF COLORADO
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-22 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10316936
关键词：
Abbreviations Address Automobile Driving Binding Binding Proteins Biochemical Biological Biology Cell physiology Cells Chromatin Code Complex Consensus Crystallization Cues DNA Binding DNA Binding Domain Data Data Set Databases Differentiation and Growth Double-Stranded RNA EMSA Electrophoretic Mobility Shift Assay Elements Enhancers Estrogen Receptors Exhibits Family Fingers Fluorescence Anisotropy Gene Expression Gene Expression Profile Gene Expression Regulation Genes Genetic Enhancer Element Genetic Transcription Glucocorticoid Receptor Glucocorticoids Goals Grant HMGB Proteins Hormones In Vitro Investigation Life Measures Mediating Modeling Molecular Molecular Conformation Motivation Mus Nature Nuclear Nuclear Hormone Receptors Nucleic Acid Regulatory Sequences Play Prevalence Process Promoter Regions Proteins RNA RNA Binding RNA Recognition Motif RNA-Protein Interaction Regulation Reporting Research Resolution Response Elements Role Site Specificity Structure Testing Thiouridine Transcriptional Regulation Untranslated RNA base design embryonic stem cell experimental study genome-wide in vitro activity in vivo insight member mutant new therapeutic target novel nucleic acid structure pluripotency factor programs promoter receptor binding response transcription factor transcriptome ultraviolet

项目摘要

Project Summary/Abstract Nimble and responsive transcriptional control is central to all processes of life, and hence this process is regulated at a number of levels. During the last grant period, we discovered a robust, and unexpected, RNA- binding activity inherent in representative proteins from two major classes of transcriptional factors (TFs): the pluripotency factor Sox2 of the high mobility group box (HMGB) family and the glucocorticoid nuclear hormone receptor (GR). Their RNA-binding activity was found to be mediated by their DNA-binding domains and directly compete with DNA binding to their respective promoter or enhancer sequences. Furthermore, we found that this activity was structure-specific rather than sequence specific in vitro. Both TFs strongly disfavored binding ssRNA. Instead, the pluripotency factor Sox2 bound dsRNA regions while GR bound exclusively to RNA hairpin structures. These observations raise critical questions regarding the role of TF RNA-binding activity in transcriptional regulation and provide the direct motivation for the research program described here. Our corroborating discovery of RNA association of Sox2 in mouse embryonic stem cells confirms that direct RNA binding occurs in vivo. The next steps are to further define and understand the specificity for the in vivo RNA targets and determine the impact TF RNA interactions has on the transcriptional program. This includes refining our understanding of what drives this RNA binding and determine how pervasive the activity is in other TFs. We have developed an integrated and strategic research program to achieve these goals. Aim 1 capitalizes on our observation that Sox2 directly interacts with RNAs in cells and develops this critical line of inquiry in GR to develop a comprehensive RNA interactome including eRNAs. We will investigate whether transcription factors use their RNA-binding activity to provide an alternate chromatin association strategy that impacts localization and gene regulation. Finally, we take advantage of our biochemical insights to design and validate DNA- and RNA-binding separation-of-function mutants, which will allow us to directly measure the impact of loss of RNA binding on the transcriptome. In Aim 2, we turn to understanding the molecular nature of RNA association to Sox2 and GR, first through the use of eCLIP strategies to identify consensus binding motifs and second through high resolution structure determination of TF/RNA complexes. In Aim 3 of this program, we will establish the generality of these observations to other important transcription factors, both within and beyond the HMGB and nuclear hormone receptor families, as represented by Sox2 and GR, respectively. Together, this proposal describes a comprehensive program seeking to determine the extent and role of RNA binding in modulating the transcriptional program through interaction with classic transcription factors. The impact of this program is high because understanding the roles of genome-wide pervasive transcription and how this activity influences diverse cellular processes remains a major unanswered question in biology.

项目总结/文摘