Epigenetic Regulation of Lysosomal Ceramide Signaling and Function in Arterial Myocytes: Role of Kmt6 Gene

动脉肌细胞溶酶体神经酰胺信号和功能的表观遗传调控：Kmt6 基因的作用

基本信息

批准号：
10298620
负责人：
PinLan Li
金额：
$ 62.89万
依托单位：
UNIVERSITY OF HOUSTON
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-04-01 至 2025-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10298620
关键词：
Abbreviations Aging Agonist Applications Grants Arteries Atherosclerosis Binding Blood Vessels Calcium Calmodulin Cathepsins B Cell membrane Cells Ceramides Chronic Kidney Failure Code Collagen DNA DNA Methylation DNA Modification Methylases Defect Deposition Development Diabetes Mellitus Disease EZH2 gene Early Endosome Enzymes Epigenetic Process Extracellular Matrix Face Functional disorder Gene Deletion Gene Expression Genes Genetic Transcription Histone-Lysine N-Methyltransferase Histones Hypertension Image Knock-out Knockout Mice Lead Link Lipids Lysine Lysosomes Medial Mediating Membrane Proteins Metabolism Methylation Methyltransferase MicroRNAs Molecular Multivesicular Body Mus Muscle Cells Pathologic Pathologic Processes Patients Phenotype Play Process Production Proteins Regulation Reporting Research Role Signal Transduction Smooth Muscle Smooth Muscle Myocytes Sphingolipids Sphingomyelinase Sphingomyelins Sphingosine Stimulus Testing Time Vascular calcification Vesicle Vitamin D acid sphingomyelinase arterial stiffness base calcification cardiovascular disorder prevention cardiovascular disorder risk cardiovascular risk factor enzyme activity epigenetic regulation exosome galactosylgalactosylglucosylceramidase gene repression histone methylation interstitial late endosome mineralization new therapeutic target novel osteogenic particle patch clamp prevent programs receptor sensor sphingosine 1-phosphate trafficking treatment strategy

项目摘要

Project Summary Arterial medial calcification (AMC) and arterial stiffening are a prevalent pathological process in different pathological conditions or diseases such as hypertension, aging, atherosclerosis, diabetes and chronic kidney disease. Enhanced exosome secretion by smooth muscle cells (SMCs) has been reported to be an essential mechanism for calcifying nidus formation and extracellular matrix mineralization in the arterial wall to result in AMC. Recent studies have also shown that lysosome function plays a critical role in controlling multivesicular body (MVB) fate and enhancing exosome secretion and thereby in the development of arterial calcification. However, it remains poorly understood how lysosome function is controlled to determine exosome secretion and thereby lead to AMC. This proposal seeks to explore a novel epigenetic mechanism that regulates lysosome trafficking and exosome secretion, which may contribute to the development of AMC. This epigenetic regulation of lysosome function may be associated with the lysine methyltransferase Kmt6- mediated repression of gene transcription of Smpd1, a lysosome enzyme that hydrolyzes sphingomyelin into ceramide. Kmt6 is considered as a crucial epigenetic regulator that represses the target gene expression by methylation of lysine residue in histone proteins. In preliminary studies, we demonstrated that SMC-specific Kmt6 gene deletion exacerbated AMC and arterial stiffening, which were associated with increased Smpd1 expression and ceramide production, reduced lysosome TRPML1 channel activity, and lysosome trafficking dysfunction. These observations led us to hypothesize that Kmt6 is an essential epigenetic regulatory enzyme that controls lysosomal Smpd1-mediated sphingolipid metabolism and thereby regulates lysosome trafficking or its fusion to MVBs and subsequent exosome secretion in SMCs. Kmt6 gene defect or functional deficiency may disturb lysosome-mediated degradation of MVBs leading to increased exosome secretion, calcifying nidus formation, osteogenic transition, and ultimately AMC in face of different pathological challenges. To test this hypothesis, the following Specific Aims are proposed. Aim 1 will determine loss of Kmt6 contributes to osteogenic transition and AMC in SMC-specific Kmt6 knockout mice with analysis of SMC phenotypes and calcification. Aim 2 attempts to test whether Kmt6-mediated epigenetic regulation of Smpd1 critically controls lysosome trafficking and exosome secretion by increasing TRPML1 channel activity and associated Ca2+ release using patch clamping of isolated lysosomes and lysosome-specific Ca2+ imaging. Aim 3 will explore the molecular mechanisms how Smpd1 gene is epigenetically regulated by Kmt6 with a focus on its action on histone and DNA methylation in cultured arterial SMCs. Our findings will for the first time define an epigenetic mechanism controlling Smpd1 expression and activity via Kmt6 in SMCs and reveal a novel role of epigenetic dysregulation of sphingolipid metabolism in the development of AMC and arterial stiffening. These findings may also identify novel therapeutic targets for the treatment of AMC under different pathological conditions.

项目概要动脉内侧钙化（AMC）和动脉硬化是不同疾病中普遍存在的病理过程。病理状况或疾病，例如高血压、衰老、动脉粥样硬化、糖尿病和慢性肾病疾病。据报道，平滑肌细胞（SMC）增强的外泌体分泌是必不可少的动脉壁钙化巢形成和细胞外基质矿化的机制 AMC。最近的研究还表明，溶酶体功能在控制多囊泡中发挥着关键作用。体（MVB）命运并增强外泌体分泌，从而参与动脉钙化的发展。然而，人们对如何控制溶酶体功能以确定外泌体的分泌仍知之甚少。从而导致AMC。该提案旨在探索一种新的表观遗传机制来调节溶酶体运输和外泌体分泌，可能有助于 AMC 的发展。这溶酶体功能的表观遗传调控可能与赖氨酸甲基转移酶 Kmt6-有关介导 Smpd1 基因转录的抑制，Smpd1 是一种溶酶体酶，可将鞘磷脂水解成神经酰胺。 Kmt6 被认为是重要的表观遗传调节因子，通过以下方式抑制靶基因表达：组蛋白中赖氨酸残基的甲基化。在初步研究中，我们证明了 SMC 特异性 Kmt6 基因缺失加剧了 AMC 和动脉硬化，这与 Smpd1 增加有关表达和神经酰胺产生、溶酶体 TRPML1 通道活性降低和溶酶体运输功能障碍。这些观察结果使我们推测 Kmt6 是一种重要的表观遗传调节酶控制溶酶体 Smpd1 介导的鞘脂代谢，从而调节溶酶体运输或其与 MVB 的融合以及随后在 SMC 中分泌的外泌体。 Kmt6 基因缺陷或功能缺陷可能会干扰溶酶体介导的 MVB 降解，导致外泌体分泌增加、钙化病灶形成、成骨转变以及最终AMC面临不同的病理挑战。测试根据这一假设，提出以下具体目标。目标 1 将确定 Kmt6 的损失会导致 SMC 特异性 Kmt6 敲除小鼠的成骨转变和 AMC，并分析 SMC 表型和钙化。目标 2 尝试测试 Kmt6 介导的 Smpd1 表观遗传调控是否关键控制通过增加 TRPML1 通道活性和相关 Ca2+ 来促进溶酶体运输和外泌体分泌使用分离的溶酶体的膜片钳和溶酶体特异性 Ca2+ 成像来释放。目标3将探索 Smpd1 基因如何受 Kmt6 表观遗传调控的分子机制，重点关注其对培养的动脉 SMC 中的组蛋白和 DNA 甲基化。我们的研究结果将首次定义表观遗传学通过 Kmt6 在 SMC 中控制 Smpd1 表达和活性的机制，揭示了表观遗传的新作用 AMC 发展和动脉硬化过程中鞘脂代谢失调。这些发现还可以确定在不同病理条件下治疗 AMC 的新治疗靶点。