Alternative Protein Isoforms in Ventricular Remodeling

心室重构中的替代蛋白质亚型

基本信息

批准号：
10660087
负责人：
Maggie Lam
金额：
$ 38.88万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-06-01 至 2027-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10660087
关键词：
Address Affect Aging Alternative Splicing Architecture Area Biological Assay Biological Process Biophysics Cardiac Complement Computing Methodologies Custom Data Development Disease Etiology Exons Failure Functional disorder Gene Expression Genes Genetic Goals Guide RNA Half-Life Heart Heart Diseases Heart Hypertrophy Heart failure Introns Knowledge Large-Scale Sequencing Mass Spectrum Analysis Mediating Metabolic Methods Modeling Molecular Mus Nonsense-Mediated Decay Pathologic Pathway interactions Pattern Phenotype Post-Translational Protein Processing Protein Databases Protein Isoforms Proteins Proteome Proteomics Quality Control RNA Splicing RNA-Binding Proteins Regulation Role Solubility Stable Isotope Labeling Structure System Tissue-Specific Splicing Tissues Transcript Translating Variant Ventricular Remodeling biophysical techniques deep learning detection assay experimental study gene regulatory network in vivo insight mouse model multiple omics prediction algorithm programs protein folding protein function protein structure response tool transcriptome sequencing

项目摘要

PROJECT SUMMARY Cardiac hypertrophy and failure involve a rewiring of gene expression through alternative splicing, but it remains unclear which splicing changes are relevant to disease development, and how splice variants affect protein function. Although many alternative transcript isoforms have been discovered, not all are translated into proteins and instead may be degraded via non-sense mediated decay or co-translational protein quality control. Proteomics methods that can identify and quantitate splice variant proteins empirically and on a large scale provide essential tools to study how alternative splicing regulates cardiac gene expression. We and others showed that tissue-specific splice variant proteins may be identified using a combined RNA sequencing and mass spectrometry approach. Accordingly, our goal here is to examine the mechanisms by which alternative splicing regulates the genetic program in hypertrophic and failing hearts, by identifying the proteins and pathways that are coordinately regulated by splicing, the resulting complement of protein isoform species (`proteoforms') in the heart, and the consequences of proteoform sequences on protein structure and function. Specifically, we plan to: (1) apply a quantitative RNA-guided proteomics framework to identify key isoform switches at the transcript and the protein level, with emphasis on the changes in splice factors and RNA-binding proteins in mouse models of systolic and diastolic dysfunction; (2) combine RNA-guided proteomics and proteome-wide biophysics approaches, we will interrogate the impact of splice variants on protein structure and thermal stability, and discover significant isoforms through alternative splicing, intrinsically disordered, and regulatory post-translational modification modules. We anticipate the successful completion of these aims will generate new conceptual insights into how alternative splicing regulatory networks reprogram the cardiac proteome in pathological remodeling and heart failure, and more generally, contribute to methods and concepts to elucidate the regulation and function of alternative splicing in the heart.

项目摘要心脏肥大和衰竭涉及通过替代剪接重新布线基因表达，但仍保持不清楚哪些剪接变化与疾病的发展有关，以及剪接变体如何影响蛋白质功能。尽管已经发现了许多替代成绩单同工型，但并非全部转化为蛋白质和蛋白质可能通过非义介导的衰减或共同翻译蛋白质质量控制降解。蛋白质组学方法可以通过经验和大规模识别和量化剪接变体蛋白提供基本工具来研究替代剪接如何调节心脏基因表达。我们和其他人表明可以使用组合的RNA测序鉴定组织特异性的剪接变异蛋白和质谱法。因此，我们的目标是检查替代方案的机制通过识别蛋白质和通过剪接协调调节的途径，蛋白质同工型的补体（“蛋白质成型”）在心脏中，以及蛋白质序列对蛋白质结构和功能的后果。具体而言，我们计划：（1）应用定量RNA引导的蛋白质组学框架来识别关键同工型转录本和蛋白质水平的开关，重点是剪接因子和RNA结合的变化收缩和舒张功能障碍的小鼠模型中的蛋白质；（2）结合RNA引导的蛋白质组学和全蛋白质组生物物理学方法，我们将询问剪接变体对蛋白质结构的影响和热稳定性，并通过替代剪接，本质上无序和调节后翻译修饰模块。我们预计这些目标将成功完成生成有关替代剪接调节网络如何重新编程心脏的新概念见解病理重塑和心力衰竭的蛋白质组，更普遍地有助于方法和概念阐明心脏中替代剪接的调节和功能。