Trimming the fat with small proteins: Micropeptides in adipogenesis

用小蛋白质减少脂肪：脂肪生成中的微肽

基本信息

批准号：
10655394
负责人：
Victor Jieh Pong Pai
金额：
$ 7.43万
依托单位：
SALK INSTITUTE FOR BIOLOGICAL STUDIES
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-01 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10655394
关键词：
3T3-L1 Cells Address Adipocytes Adipose tissue Adult Affect Alanine Algorithms Anti-Obesity Agents Antibodies Biological Biology Blood Glucose Body Weight Body Weight decreased CRISPR library CRISPR screen CRISPR/Cas technology Cell Line Cells Code Codon Nucleotides Computational algorithm DNA sequencing Data Databases Drug Targeting Energy Intake Epidemic FDA approved Fatty Liver Fatty acid glycerol esters Generations Genes Genetic Genetic Variation Genome Genomics Goals Homeostasis Human Genome Hypertrophy Immunofluorescence Immunologic Immunoprecipitation Individual Inflammation Insulin Resistance Knock-out Knowledge Laboratories Liver Location Mass Spectrum Analysis Metabolic Diseases Methods Molecular Biology Monitor Mouse Strains Mus Mutagenesis Non-Insulin-Dependent Diabetes Mellitus Nonesterified Fatty Acids Obesity Open Reading Frames Output Overweight Pathogenesis Pathway interactions Patients Peptides Phenotype Phosphorylation Physiological Physiology Play Proteins Proteomics Publishing Retinal blind spot Role Scanning Scientist Serine Signal Transduction Technology Testing Translating Triglycerides Validation Work adipocyte biology adipocyte differentiation candidate validation cohort diverse data effective therapy experience experimental study glucose uptake in vitro Model insulin signaling interest lipid biosynthesis live cell imaging mouse genome mouse model multiple omics new therapeutic target next generation novel obesity treatment overexpression protein protein interaction ribosome profiling transcriptome sequencing

项目摘要

Project Summary/Abstract Obesity is an ongoing epidemic, with more than two-thirds of adults in the US considered to be overweight or obese. Treatment options for these patients, including all FDA approved anti-obesity drugs, are ineffective for long term weight loss. The lack of effective treatments for obesity signals a gap in knowledge. The white adipose tissue is responsible for maintaining energy storage and release to maintain fuel homeostasis in the body. Disruption of white adipocyte function due to excess energy intake is a major contributor to the pathogenesis of obesity, resulting in insulin resistance, type 2 diabetes, and other metabolic diseases. Recent advancements in gene sequencing and mass spectrometry technology have allowed scientists to identify small Open Reading Frames (smORFs), which encodes functional microproteins that have a wide range of biological roles. The Saghatelian laboratory has pioneered a suite of multi-omic platforms that integrate proteomics, ribosome profiling (Ribo-Seq), and RNA sequencing to accurately annotate smORFs of less than 150 codons. By performing Ribo- Seq on models of murine adipocytes, the applicant has identified 1721 novel smORFs in the mouse genome. The great majority of these smORFs are uncharacterized. This major goal in this proposal is to address this knowledge gap by characterizing novel adipocyte smORFs that are involved in adipogenesis (Aim 1) and insulin signaling (Aim 2). Using a CRISPR/Cas9 library screen specifically targeting the 1721 novel smORFs, the applicant has already identified ~20 smORFs that specifically inhibit adipogenesis. Subaim 1.1 will validate these findings through generation of smORF knockout 3T3-L1 preadipocytes to assess how specific smORFs affect adipocyte differentiation, followed by rescue experiments through overexpressing smORFs in knockout cells. In Subaim 1.2, the applicant will dive deep into the function of a specific microprotein identified in Subaim 1.1 by characterizing its cellular location and microprotein-protein interaction. In Aim 2, the applicant will exploit the genomic and physiology data from the Diversity Outbred (DO) mice to identify smORFs that correlates with physiological changes, such as body weight, blood glucose, and triglycerides. The applicant will determine whether knocking out the smORFs identified from this analysis in 3T3-L1 cells will affect insulin signaling and glucose uptake under normal conditions and in in vitro models of insulin resistance. Completion of these studies will reveal how smORF and microproteins regulate adipocyte differentiation and insulin signaling.

项目摘要/摘要肥胖是一种持续的流行病，在美国，有超过三分之二的成年人被认为是超重或肥胖。这些患者的治疗选择，包括所有FDA批准的抗肥胖药物，无效长期体重减轻。缺乏对肥胖症的有效治疗信号是知识的差距。白脂肪组织负责保持能量存储并释放以维持体内燃料稳态。由于摄入过多而导致的白色脂肪细胞功能的破坏是导致发病机理的主要原因肥胖，导致胰岛素抵抗，2型糖尿病和其他代谢疾病。最近的进步基因测序和质谱技术使科学家可以识别出小的开放阅读框架（SMORF），它编码具有广泛生物学作用的功能性微蛋白。这 Saghatelian实验室已经开创了一套多摩变平台，这些平台整合了蛋白质组学，核糖体谱分析（Ribo-Seq）和RNA测序以准确注释少于150个密码子的SMORF。通过执行肋骨 SEQ在鼠脂肪细胞模型上，申请人在小鼠基因组中确定了1721个新型SMORF。这些smorfs中的绝大多数是未经表征的。该提案中的主要目标是解决这个问题通过表征参与脂肪生成（AIM 1）和胰岛素的新型脂肪细胞Smorfs来表征知识差距信号传导（AIM 2）。使用CRISPR/CAS9库屏幕专门针对1721个小说Smorfs 申请人已经确定了约有20种特异性抑制脂肪形成的smorf。 Subaim 1.1将验证这些发现通过产生SMORF敲除3T3-L1前脂细胞来评估特定SMORF的方式影响脂肪细胞的分化，然后通过过表达敲除中的smorfs进行救援实验细胞。在Subaim 1.2中，申请人将深入研究Subaim中鉴定的特定微蛋白的功能 1.1通过表征其细胞位置和微蛋白 - 蛋白质相互作用。在AIM 2中，申请人将利用来自多样性杂种（DO）小鼠的基因组和生理数据，以识别与生理变化，例如体重，血糖和甘油三酸酯。申请人将确定在3T3-L1细胞中从该分析中鉴定出的SMORF是否会影响胰岛素信号传导和在正常条件下和胰岛素抵抗的体外模型中葡萄糖摄取。这些研究的完成将揭示SMORF和微蛋白如何调节脂肪细胞分化和胰岛素信号传导。