Interrogating the role of m6A mRNA methylation in the aging of the β-cell and diabetes

探讨 m6A mRNA 甲基化在 β 细胞衰老和糖尿病中的作用

• 批准号: 10644215
• 负责人: Dario F De Jesus
• 金额: $ 9万
• 依托单位: JOSLIN DIABETES CENTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10644215
• 关键词: ATM activation; Acceleration; Adult; Age; Aging; Area; Autoimmune; Automobile Driving; Beta Cell; Bioinformatics; Biology of Aging; Candidate Disease Gene; Cell Aging; Cell Line; Cell Survival; Cell physiology; Cell secretion; Chemicals; Complement; DNA Damage; Data; Data Analyses; Data Set; Dependovirus; Development; Diabetes Mellitus; Disease; Exhibits; Female; Foundations; Future; Gene Expression; Genes; Genetic Models; Goals; Health; High Fat Diet; Homologous Gene; Human; Immunoprecipitation; Impairment; Inbred NOD Mice; Inflammation; Insulin Resistance; Knockout Mice; Messenger RNA; Metabolism; Methods; Methylation; Methyltransferase; Modification; Molecular; Molecular Biology; Molecular Target; Mus; Non-Insulin-Dependent Diabetes Mellitus; Pathway interactions; Patients; Phase; Physiology; Publishing; RNA; RNA Splicing; Regulation; Research; Research Personnel; Ribonucleosides; Role; Testing; Time; Training; Transcript; Up-Regulation; Work; aged; beta-Galactosidase; candidate validation; career; comparison control; crosslink; crosslinking and immunoprecipitation sequencing; experience; experimental study; functional decline; improved; in vivo; in vivo evaluation; insight; islet; mRNA Translation; methylome; mouse genetics; novel; novel therapeutics; protein complex; response; senescence; single-cell RNA sequencing; stressor; therapeutic development; therapeutic target; transcriptomics; type I and type II diabetes; ATM activation; Acceleration; Adult; Age; Aging; Area; Autoimmune; Automobile Driving; Beta Cell; Bioinformatics; Biology of Aging; Candidate Disease Gene; Cell Aging; Cell Line; Cell Survival; Cell physiology; Cell secretion; Chemicals; Complement; DNA Damage; Data; Data Analyses; Data Set; Dependovirus; Development; Diabetes Mellitus; Disease; Exhibits; Female; Foundations; Future; Gene Expression; Genes; Genetic Models; Goals; Health; High Fat Diet; Homologous Gene; Human; Immunoprecipitation; Impairment; Inbred NOD Mice; Inflammation; Insulin Resistance; Knockout Mice; Messenger RNA; Metabolism; Methods; Methylation; Methyltransferase; Modification; Molecular; Molecular Biology; Molecular Target; Mus; Non-Insulin-Dependent Diabetes Mellitus; Pathway interactions; Patients; Phase; Physiology; Publishing; RNA; RNA Splicing; Regulation; Research; Research Personnel; Ribonucleosides; Role; Testing; Time; Training; Transcript; Up-Regulation; Work; aged; beta-Galactosidase; candidate validation; career; comparison control; crosslink; crosslinking and immunoprecipitation sequencing; experience; experimental study; functional decline; improved; in vivo; in vivo evaluation; insight; islet; mRNA Translation; methylome; mouse genetics; novel; novel therapeutics; protein complex; response; senescence; single-cell RNA sequencing; stressor; therapeutic development; therapeutic target; transcriptomics; type I and type II diabetes

PROJECT SUMMARY Although the etiopathogenesis of Type 1 (T1D) and Type 2 diabetes (T2D) is different, decreased functional β-cell mass is a central feature in both diseases. Concerted research efforts aim at developing strategies to improve β-cell survival and restore β-cell function in patients with diabetes. Cellular senescence is a major hallmark of aging. Recent work has demonstrated that β-cell senescence is a common contributor to Type 1 diabetes (T1D) and Type 2 diabetes (T2D). RNA modifications are emerging as important modulators of many hallmarks of aging. We have discovered that m6A mRNA methylation is essential for β-cell function and survival, and recently characterized the m6A landscape of human T1D and T2D islets. Our preliminary data shows that the m6A eraser ALKBH5 is upregulated in aged human β-cells and differentially m6A methylated genes in T1D and T2D are enriched for DNA damage response and senescence pathways compared to controls. The goal of this application is to use state-of-the art methods to characterize the m6A methylome of the aging β- cell and identify m6A-regulated pathways that underlie the accelerated β-cell senescence in diabetes. I will test the overarching hypothesis that aging β-cells exhibit transcript-specific m6A hypomethylation and consequent upregulation of mRNAs involved in driving cell senescence, and that m6A is therefore a valuable therapeutic target for diabetes. In Specific Aim 1, I will characterize the temporal m6A landscape of the aging β-cell. Completion of Aim 1, which will take place during K99 phase, will provide me with training in aspects of aging biology and further experience with bioinformatics analysis of m6A data necessary to independently complete the R00 phase and future R01 submissions. In Specific Aim 2, which will span the K99 and R00 phases, I will identify the molecular targets of ALKBH5 in the human β-cell transition to senescence. Completion of the sub- aim of Aim 2 proposed during the K99 phase will provide me with the training in photoactivatable ribonucleoside- enhanced crosslinking and immunoprecipitation (PAR-CLIP) sequencing and data analysis necessary to complement Aim 1, and independently complete Aim 2 during the R00 phase. For Specific Aim 3, I will target m6A levels to reduce β-cell senescence and improve diabetes in vivo. Completion of this aim will allow me to validate candidate genes identified in Aim’s 1 and 2 and to test in vivo the role of Alkbh5 in driving the accelerated senescence seen in β-cells in diabetes. These experiments are novel as they combine omics-based approaches, mouse genetic models, physiology, and molecular biology to examine the mechanistic role of m6A, and particularly ALKBH5, in β-cell senescence transition in diabetes.

项目摘要 尽管1型（T1D）和2型糖尿病（T2D）的疗法发生不同，但精致 功能性β细胞质量是两种疾病的核心特征。一致的研究工作旨在发展 改善糖尿病患者的β细胞存活和恢复β细胞功能的策略。细胞感应是 衰老的主要标志。最近的工作表明，β细胞的感应是 1型糖尿病（T1D）和2型糖尿病（T2D）。 RNA修饰是作为重要调节剂的出现 衰老的许多标志。我们发现M6a mRNA甲基化对于β细胞功能至关重要 生存，最近描述了人类T1D和T2D小岛的M6A景观。我们的初步数据 表明M6A橡皮擦ALKBH5在老年人的β细胞中进行了更新，M6A甲基化的 与对照组相比，T1D和T2D中的基因富含DNA损伤响应和感应途径。 该应用的目的是使用最先进的方法来表征衰老β-的M6A甲基团 细胞并确定糖尿病中加速β细胞感应的M6A调节途径。我会测试 总体假设，即衰老的β细胞暴露了转录本特异性的M6a甲基化及其因此 因此，涉及驱动细胞感应的mRNA的上调，因此M6A是一种有价值的疗法 糖尿病的靶标。在特定的目标1中，我将表征衰老β细胞的临时M6A景观。 AIM 1的完成将在K99阶段进行，将为我提供衰老方面的培训 生物学和进一步的经验，对独立完成所需的M6A数据的生物信息学分析 R00阶段和未来R01提交。在跨越K99和R00阶段的特定目标2中，我将 鉴定人β细胞转变为敏感的AlkBH5的分子靶标。完成子的完成 在K99阶段提出的AIM 2的目标将为我提供光活化核苷的训练。 增强的交联和免疫沉淀（PAR-CLIP）测序和数据分析所必需的 补充目标1，并在R00阶段独立完成AIM 2。对于特定的目标3，我将针对 M6a水平以降低β细胞感应并改善体内糖尿病。这个目标的完成将使我能够 在AIM的1和2中鉴定的验证候选基因，并在体内测试ALKBH5在驱动加速的作用 在糖尿病的β细胞中看到的衰老。这些实验是新颖的，因为它们结合了基于OMICS的方法， 小鼠遗传模型，生理学和分子生物学，以检查M6A的机理作用， 特别是ALKBH5，在糖尿病中的β细胞感应过渡中。