Macronutrient Regulation of Alternative Pre-mRNA Splicing

替代性前 mRNA 剪接的常量营养素调控

• 批准号: 8418354
• 负责人: Scot R Kimball
• 金额: $ 15.3万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-03-20 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8418354
• 关键词: Address; Affect; Alternative Splicing; Amino Acids; Attenuated; Biological Markers; Body Composition; Body Weight; Carbohydrates; Cell Culture Techniques; Cell Line; Consumption; Diet; Dietary Fatty Acid; Disease; Energy Metabolism; Excision; Exhibits; Exons; Experimental Models; Fatty Acids; Fatty acid glycerol esters; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Genes; Genetic Transcription; Genetic Translation; Goals; Homeostasis; Impairment; Inflammation; Inflammatory; Inflammatory Response; Knowledge; Lead; Ligation; Lipids; Macronutrients Nutrition; Mammalian Cell; Mammals; Mediating; Messenger RNA; Metabolic; Metabolic Diseases; Metabolic syndrome; Metabolism; Modification; Molecular; Mono-S; Moths; Muscle; Non obese; Nutrient; Obesity; Organism; Pathway interactions; Pattern; Performance; Phosphorylation; Plasma; Play; Prevention approach; Process; Production; Protein Isoforms; Proteins; RNA Splicing; Rattus; Regulation; Research; Role; Saturated Fatty Acids; Signal Pathway; Signal Transduction; Suggestion; System; Tissue Sample; Troponin T; Unsaturated Fats; Variant; Weight; Zucker Rats; base; cytokine; feeding; innovation; mRNA Precursor; novel; polyunsaturated fat; protein expression; protein function; research study; response

DESCRIPTION (provided by applicant): We have recently discovered a novel and evolutionarily conserved homeostatic response wherein alternative splicing of the pre-mRNA encoding troponin T, a gene that affects muscle force production, is tightly regulated in response to changes in body weight. The effect is based on weight rather than mass or compartmentation of mass within the body because an external load has the same effect as an increment in native body weight. However, this response occurs differently if the load comprises fat, as load-induced changes in troponin T pre-mRNA alternative splicing are impaired in obese, but not lean, Zucker rats, leading to inappropriate expression of particular troponin T isoforms. In preliminary studies, we found a similar dysregulation in rats fed a high-fat diet enriched in saturated fatty acids. Notably, high-fat diet-induced changes in troponin T mRNA alternative splicing manifest prior to detectable alterations in either body weight or body composition, suggesting that alternative splicing is directly modulated in response to the fat content of the diet. Hence, we seek to characterize and examine experimentally the mechanisms whereby a high-fat diet causes deviation from the normal weight response in troponin T alternative splicing. We hypothesize that the difference is due to the known inflammatory effects of a high-fat diet and so by studying this system we seek to answer important questions on a number of levels via the following three specific aims: (1) assess the effects of dietary fatty acids, e.g. saturated vs. unsaturated, on quantitative alternative splicing of pre-mRNA at the level of our focal gene and across the transcriptome, as revealed by an exon array, (2) assess the effects of a high-fat diet on alterations in the signaling pathways and regulatory mechanisms that control quantitative alternative splicing of pre-mRNA, and, (3) establish a cell culture experimental model based on cytokine and lipidomic analysis of plasma and tissue samples to define signaling pathways and molecular mechanisms through which selected nutrients act to control alternative splicing. From these experiments, we will obtain an unprecedented scale and depth of understanding of how quantitative variation in alternative splicing is controlled, and how diet affects that regulation. In addition, these results will open a new window into how diet changes homeostative pathways involved in body weight homeostasis. Overall, the studies proposed here are highly original and will address a deficit in our knowledge about the plasticity of quantitative alternative splicing in general, and mechanisms through which macronutrients affect and in some cases disrupt the way metazoans functionally and metabolically adapt to changes in their weight. We expect the proposed research to reveal biomarkers for pre-disease states caused by poor diet, and candidate molecules and pathways for pharmacological manipulation to provide new and innovative approaches for the prevention and treatment of metabolic disorders.

描述（由申请人提供）：我们最近发现了一种新的和进化上保守的稳态应答，其中编码肌钙蛋白T的前体mRNA（一种影响肌肉力量产生的基因）的选择性剪接响应于体重变化而受到严格调控。该效应基于体重，而不是体内的质量或质量区室，因为外部负荷与天然体重的增加具有相同的效应。然而，如果负荷包括脂肪，则这种反应发生不同，因为负荷诱导的肌钙蛋白T前mRNA选择性剪接的变化在肥胖但不瘦的Zucker大鼠中受损，导致特定肌钙蛋白T亚型的不适当表达。在初步研究中，我们在喂食富含饱和脂肪酸的高脂肪饮食的大鼠中发现了类似的失调。值得注意的是，高脂饮食诱导的肌钙蛋白T mRNA选择性剪接的变化在体重或身体组成的可检测变化之前就表现出来，这表明选择性剪接是直接调节饮食中的脂肪含量的。因此，我们试图表征和实验研究的机制，即高脂肪饮食导致偏离正常体重反应的肌钙蛋白T选择性剪接。我们假设这种差异是由于高脂肪饮食的已知炎症效应造成的，因此通过研究这个系统，我们试图通过以下三个具体目标来回答一些层面上的重要问题：（1）评估膳食脂肪酸（例如饱和脂肪酸与不饱和脂肪酸）对在我们的焦点基因水平上和跨转录组的前mRNA的定量可变剪接的影响，如外显子阵列所揭示的，（2）评估高脂肪饮食对控制前体mRNA的定量可变剪接的信号传导途径和调节机制的改变的影响，以及，（三）建立基于血浆和组织样品的细胞因子和脂质组学分析的细胞培养实验模型，以确定信号传导途径和分子机制，所选营养素通过所述信号传导途径和分子机制起作用，控制可变剪接。从这些实验中，我们将获得前所未有的规模和深度的理解，了解选择性剪接的数量变化是如何控制的，以及饮食如何影响这种调节。此外，这些结果将打开一个新的窗口，了解饮食如何改变参与体重稳态的稳态途径。总的来说，这里提出的研究是非常原始的，并将解决我们对定量选择性剪接的可塑性的认识不足，以及大量营养素影响和在某些情况下破坏后生动物功能和代谢适应体重变化的机制。我们希望拟议的研究能够揭示由不良饮食引起的疾病前状态的生物标志物，以及用于药理学操作的候选分子和途径，为预防和治疗代谢紊乱提供新的创新方法。

项目成果

Effects of age and hindlimb immobilization and remobilization on fast troponin T precursor mRNA alternative splicing in rat gastrocnemius muscle.

• DOI: 10.1139/apnm-2015-0381
• 发表时间: 2016-02
• 期刊: Applied physiology, nutrition, and metabolism = Physiologie appliquee, nutrition et metabolisme
• 作者: Ravi S;Schilder RJ;Berg AS;Kimball SR
• 通讯作者: Kimball SR