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Mechanisms of RNA localization and translational regulation on the endoplasmic reticulum

内质网RNA定位和翻译调控机制

基本信息

批准号：
10460908
负责人：
Christopher V. Nicchitta
金额：
$ 64.66万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-06 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10460908
关键词：
Address Aging Binding Proteins Biochemical Biogenesis Biological Biology Cell Communication Cell Culture System Cell surface Cells Code Development Disease Endoplasmic Reticulum Gene Expression Genes Genetic Transcription Hormone secretion Individual Laboratories Learning Location Mammalian Cell Mediating Membrane Proteins Memory Messenger RNA Metabolism Modeling Neurotransmitters Pathway interactions Peptide Signal Sequences Process Protein Biosynthesis Protein translocation Proteins Proteome RNA RNA-Binding Proteins Regulation Research Ribosomes Role Signal Recognition Particle Signal Transduction Site System Testing Transcript Translation Initiation Translational Regulation Translations crosslinking and immunoprecipitation sequencing in vivo insight protein function proteostasis ribophorin I ribosome profiling secretory protein signal sequence receptor tissue/cell culture trafficking transcriptome transcriptome sequencing

项目摘要

The endoplasmic reticulum (ER) is the subcellular site of secretory and membrane protein synthesis and performs critical functions in secretory/membrane protein biogenesis and cellular proteostasis. In addition its established role in secretory/membrane protein synthesis, recent studies examining the mRNA transcriptomes of cytosolic and ER-bound ribosomes reveal that cytosolic protein transcripts are broadly represented on the ER, with ribosome footprinting analyses demonstrating translation of cytosolic protein mRNAs on ER-associated ribosomes. These findings identify an unexpected mRNA transcriptome-wide function for the ER in proteome expression and reopen fundamental questions regarding the mechanisms regulating mRNA localization and translation on the ER. Principally, where current models posit that mRNA localization to the ER is co-translational and signal sequence-dependent, the abundant presence and translation of cytosolic protein mRNAs on the ER indicates that either alternative and/or multiple pathways mediate mRNA localization to the ER. As well, and although SRP pathway function in protein translocation is well established, the question of SRP pathway function in mRNA localization to the ER remains largely unexplored. Also of significance, the recent findings that a number of translocon-associated proteins, including Sec61α,β, TRAPα, ribophorin I, and p180, are mRNA binding proteins (RBPs) suggest previously unappreciated roles for ER resident RBPs in the biology of RNA localization and translation on the ER. This proposal merges three primary research themes of our laboratory; SRP pathway function in mRNA and ribosome localization to the ER; ii) ER-localized translation initiation as a mechanism of localized protein synthesis, and iii) RNA binding protein function in RNA localization and translational regulation, to address new questions regarding cellular mechanisms of mRNA and ribosome localization to the ER. Building on the past decade and a half of our research into RNA localization and translational regulation on the ER, including founding evidence identifying an mRNA transcriptome-wide role for the ER in cellular proteome expression, the proposed research will utilize mammalian tissue culture cell systems, gene editing and silencing approaches, RNA-seq and Ribo-seq transcriptome analyses, HITS-CLiP and PAR-CLIP studies of ER RNA binding proteins and their RNA interactomes, and biochemical analyses of the subcellular organization of the translation machinery, to obtain new insights into the cellular organization and regulation of proteome expression. This research is expected to advance understanding into the systems and pathways governing post-transcriptional gene expression in the cell. In emphasizing in vivo analyses and native biosynthetic approaches to the study of RNA and ribosome trafficking dynamics, this research is significant in its efforts to rigorously test existing paradigms and advance understanding of cellular mechanisms of localized protein synthesis.

内质网（ER）是分泌和合成膜蛋白的亚细胞部位