The regulation of the histone code during cardiac hypertrophy

心脏肥大过程中组蛋白密码的调节

• 批准号: 10373727
• 负责人: Maha Abdellatif
• 金额: $ 66.54万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-01 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10373727
• 关键词: Acetylation; Aging; Butyryl-CoA dehydrogenase; CDK9 Protein Kinase; Carbohydrates; Cell Nucleus; Cells; ChIP-seq; Coenzyme A; Data; Diet; Dietary Fats; Disease Outcome; Disease Progression; Enzymes; Fat-Free Diets; Fatty Acids; Fatty acid glycerol esters; Gene Expression; Genes; Genetic Transcription; Genomics; Glucose; Goals; Health; Heart; Heart Hypertrophy; Heart failure; High Fat Diet; Histone Code; Histones; Human; Hypertrophy; Ketone Bodies; Ligase; Lysine; Mediating; Metabolic; Metabolism; Molecular; Mus; Nonesterified Fatty Acids; Nuclear; Outcome; Oxidoreductase; Pathologic; Regulation; Reporting; Role; Source; Stress; Sum; Transcriptional Regulation; acyl group; beta-Hydroxybutyrate; dietary; disease prognosis; genome-wide; histone methylation; histone modification; improved; improved outcome; ketogenic diet; ketogentic; knock-down; oxidation; pressure; promoter; recruit; transcriptome sequencing

Abstract Our goal is to investigate the impact of diet and pressure overload on the histone code, and how this influences changes in gene expression in the healthy and hypertrophied/failing hearts and, in turn, how it impacts progression of the disease. Deciphering the histone code and how diet can modify it, provides us an educated means to exploit it to our advantage, especially during pathological conditions. Acetylation and methylation of histone lysine (K) residues were the first histone modifications discovered and are, therefore, the most widely studied and understood. However, to-date, there are 11 confirmed modifiers of histone lysine residues, including the acyl groups butyryl (Bu), crotonyl (Cr), and b-hydroxybutyrate (bHB) 1, whose source, genomic distribution, and functional relevance, remain largely unknown in the heart, and are the focus of our study. Our recent findings uniquely show that dietary fat is a major regulator of histone butyrylation, including H3K9-butyryl (H3K9Bu). Using genome-wide chromatin immunoprecipitation-sequencing (ChIP-Seq), we show that H3K9Bu is abundant at all transcriptionally active promoters. Both a high-fat diet and stress accelerated the conversion of butyryl-CoA to crotonyl-CoA via acyl-CoA dehydrogenase short chain (ACADS), resulting in a substantial reduction in global promoter-H3K9Bu. A deletion of ACADS both in the mouse heart and in human cells reversed this effect and increased promoter and gene-body H3K9Bu. Paradoxically, though, a fat-free diet had the highest levels of H3K9Bu. Deletion of fatty acid synthetase (FASN), abolished H3K9Bu in cells maintained in a glucose-rich, fatty acid-free, but not in a fatty acid-rich, medium, proving that fatty acid synthesis from carbohydrates substitutes for dietary fat as a source butyryl-CoA. In contrast to H3K9Bu, there were minimal dietary-induced changes in H3K9-acetyl (H3K9ac) levels. Importantly, RNA-sequencing (RNA-Seq) revealed that diet-induced changes in H3K9Bu abundance in the mouse heart was associated with differential changes in gene expression, but only when stressed by pressure overload. Moreover, promoter-H3K9Bu levels inversely correlated with the extent of changes in gene expression levels, as evidenced by the more robust changes seen in the hearts of mice on a, short-term, high-fat vs a fat-free diet, as well as, after deletion of the ACADS. Interestingly, H3K9Bu abundance inversely correlated with H3K9-crotonyl (H3K9Cr) and Cdk9. In sum, our data uniquely show that H3K9Bu is enriched at active promoters, is negatively regulated by high-fat and stress in an ACADS-dependent fashion, and its abundance inversely correlates with stress-induced changes in gene expression. We are proposing that histone H3K9Bu, H3K9Cr, and H3K9-b-hydroxybutyryl (H3K9bHB), are products of the b-oxidation intermediates, butyryl-CoA, crotonyl-CoA, and b-hydroxybutyryl-CoA, or the ketone body, b-hydroxybutyrate, which serve as substrates for histones modifications. These marks are labile and differentially influence pressure overload-induced gene expression, but not baseline expression. Specifically, as H3K9Bu decreases it is replaced by H3K9Cr during a high-fat diet. This exchange exaggerates gene expression and worsens the outcome of cardiac failure. Conversely, H3K9bHB that increases during a ketogenic diet has the opposite effect, as it is reported to have beneficial effects on health and aging. This differential influence of the histone marks on gene expression is mediated by regulating the recruitment of Cdk9 to gene promoters. We hypothesize that 1) A high-fat diet (60 Kcal% fat, 20 Kcal% carb), or pressure overload, accelerates the conversion of nuclear butyryl- CoA to crotonyl-CoA in an ACADS-dependent manner, thus, reducing H3K9Bu and increasing H3K9Cr, which is responsible for exaggerating stress-induced gene expression and worsening the outcome of heart failure (HF). In contrast, a ketogenic diet (84 Kcal% fat, 0% carb) will produce high levels of b-hydroxybutyryl that will increase H3K9bHB, which curbs changes in stress-induced gene expression in a b-hydroxybutyrate dehydrogenase (BDH1)-dependent fashion, improving the outcome of HF. Supplementing a diet with b-hydroxybutyrate will also increase H3K9bHB, with similar beneficial effects. 2) Therefore, knockdown of ACADS reduces the conversion of butyryl-CoA to crotonyl-CoA, increasing H3K9Bu and improving the outcome of heart failure during a high-fat diet. Conversely, deletion or inhibition of BDH1 reduces H3K9bHB and worsens conditions. 3) H3K9Cr enhances the dynamics of cyclin-dependent kinase 9 (Cdk9) recruitment to promoters during stress, whereas, H3K9Bu and H3K9bHB temper it, thus, reducing the extent of changes in gene expression and improving disease outcome. The specific aims are: 1) Examine the effects of high-fat, ketogenic, and b-hydroxybutyrate-enriched diets on the genome-wide distribution and changes in H3K9Bu, H3K9Cr and H3K9bHB, changes in gene expression, and their impact on the progression of cardiac hypertrophy and failure. 2) Investigate the roles of ACADS and BDH1 in regulating the levels of H3K9Bu, H3K9Cr, and H3K9bHb, and the progression of cardiac hypertrophy and failure. 3) Investigate the role of Cdk9 in mediating the differential transcriptional regulation directed by promoter-H3K9Cr vs. H3K9Bu or H3K9bHB during cardiac hypertrophy and failure.

摘要 我们的目标是研究饮食和压力超负荷对组蛋白编码的影响，以及这种影响是如何产生的。 健康和肥厚/衰竭心脏中基因表达的变化及其反过来如何影响 疾病的发展。破译组蛋白密码以及饮食如何修改它，为我们提供了一个受过教育的 意味着利用它来利用我们的优势，特别是在病态情况下。乙酰化和甲基化 组蛋白赖氨酸(K)残基是第一个被发现的组蛋白修饰，因此也是最广泛的 学习和理解。然而，到目前为止，已证实的组蛋白赖氨酸残基的修饰物有11个，包括 酰基丁酰基(Bu)、巴豆基(Cr)和b-羟基丁酸酯(BHB)1，其来源、基因组分布、 和功能相关性，在很大程度上仍不为人所知，也是我们研究的重点。我们最新的发现 独特地表明，饮食脂肪是组蛋白丁酰化的主要调节因素，包括H3K9-丁酰基(H3K9Bu)。 利用全基因组染色质免疫沉淀测序(ChIP-Seq)，我们证明了H3K9Bu是丰富的 所有转录活性的启动子。高脂饮食和压力都加速了丁酰辅酶A的转化 通过酰基辅酶A脱氢酶短链(AADS)转化为巴豆酰-辅酶A，导致全球 启动子-H3K9Bu。在小鼠心脏和人类细胞中AADS的缺失逆转了这一效应，并 增加启动子和基因体H3K9Bu。然而，矛盾的是，脱脂饮食含有最高水平的 H3K9Bu。脂肪酸合成酶(FASN)的缺失，消除了H3K9Bu在细胞中维持的富含葡萄糖、脂肪 无酸，但不是在富含脂肪酸的介质中，证明了从碳水化合物合成脂肪酸是替代品 作为丁酰辅酶A来源的膳食脂肪。与H3K9Bu相比，饮食诱导的变化很小 H3K9-乙酰(H3K9ac)水平。重要的是，RNA测序(RNA-Seq)显示，饮食诱导的变化 小鼠心脏中H3K9Bu的丰度与基因表达的差异变化有关，但只有 当压力过大时。此外，启动子-H3K9Bu水平与疾病的程度呈负相关 基因表达水平的变化，从小鼠心脏中看到的更强劲的变化就可以证明， 短期内，高脂肪饮食与脱脂饮食相比，以及ACADS基因缺失后。有趣的是，H3K9Bu丰度 与H3K9-巴豆基(H3K9Cr)和CDK9呈负相关。总而言之，我们的数据唯一地表明H3K9Bu是 富含活性启动子，以Acads依赖的方式受到高脂肪和应激的负面调节， 而且它的丰度与压力诱导的基因表达变化呈负相关。我们建议 组蛋白H3K9Bu、H3K9Cr和H3K9-b-羟基丁酰基(H3K9bHB)是b-氧化的产物 中间体，丁酰辅酶A，巴豆基辅酶A和b-羟基丁酰辅酶A，或酮体，b-羟基丁酸酯， 它们作为组蛋白修饰的底物。这些标记是不稳定的，对压力有不同的影响 超负荷诱导基因表达，但不诱导基线表达。具体地说，随着H3K9Bu的减少，它被取代 在高脂肪饮食中被H3K9Cr.这种交换夸大了基因的表达，并恶化了 心力衰竭。相反，在生酮饮食中增加的H3K9bHb具有相反的效果，因为它 据报道对健康和衰老有有益的影响。组蛋白标记对基因的这种差异影响 表达是通过调节CDK9对基因启动子的募集来调节的。我们假设1)A 高脂饮食(60KCAL%脂肪，20KCAL%碳水化合物)或压力超负荷，加速核丁酰基- CoA以Acads依赖的方式转化为巴豆酰-CoA，从而降低H3K9Bu，增加H3K9Cr， 是夸大应激诱导的基因表达和恶化心力衰竭(HF)结局的罪魁祸首。 相反，生酮饮食(84KCAL%脂肪，0%碳水化合物)会产生高水平的b-羟丁酰基，这将增加 H3K9bHB，抑制应激诱导的b-羟丁酸脱氢酶基因表达的变化 (BDH1)依赖的方式，改善心衰的结局。补充饮食中的b-羟基丁酸酯也会 增加H3K9bHb，具有类似的有益效果。2)因此，击倒AADS降低了转化率 丁酰辅酶A转变为巴豆酰辅酶A，增加H3K9Bu，改善高脂状态下心力衰竭的结局 节食。相反，BDH1的缺失或抑制会减少H3K9bHb，使病情恶化。3)H3K9Cr增强 细胞周期蛋白依赖性激酶9(CDK9)在应激过程中向启动子募集的动态，而H3K9Bu 和H3K9bHb调节它，从而减少基因表达变化的程度，改善疾病 结果。具体目标是：1)检查高脂肪、生酮和富含b-羟基丁酸的影响。 日粮对H3K9Bu、H3K9cr和H3K9bHb全基因组分布及基因变化的影响 表达，以及它们在心肌肥厚和衰竭进展中的影响。2)调查 AADS和BDH1在调节H3K9Bu、H3K9cr和H3K9bHb水平及心脏疾病进展中的作用 肥大和失败。3)研究cdk9在介导差异转录调控中的作用。 在心肌肥厚和衰竭过程中，由启动子-H3K9cr与H3K9Bu或H3K9bHb进行比较。