Gene Modulation of Acetylation Modifiers to Reveal Regulatory Links to Human Cardiac Electromechanics

乙酰化修饰剂的基因调节揭示与人类心脏机电的调节联系

• 批准号: 10677295
• 负责人: MARIA POZO
• 金额: $ 3.79万
• 依托单位: GEORGE WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10677295
• 关键词: Acetylation; Action Potentials; Acute; Arrhythmia; Autoimmune Diseases; Automobile Driving; Autophagocytosis; Behavior; Biological Assay; CRISPR interference; CRISPR-mediated transcriptional activation; Calcium; Cardiac; Cardiac Electrophysiologic Techniques; Cardiac health; Cardiotoxicity; Cardiovascular Diseases; Cardiovascular system; Cell Cycle; Cells; Complex; Computer Models; Contractile Proteins; Cytoskeletal Proteins; Data; Development; Disease model; Electrophysiology (science); Emergency Situation; Epigenetic Process; Event; Exhibits; Feedback; Fibrosis; Frequencies; Functional disorder; Gene Expression; Genes; Genetic Transcription; Guide RNA; Health; Heart; Heart Injuries; Histone Deacetylase; Histone Deacetylase Inhibitor; Homeostasis; Human; Hypertrophy; Immune System Diseases; Individual; Inflammatory Response; Intervention; Ion Channel; Ischemia; Libraries; Link; Malignant Neoplasms; Measurement; Measures; Mechanics; Mitochondria; Modeling; Morbidity - disease rate; Oncology; Optics; Oxidative Stress; Pathology; Phenotype; Predisposition; Process; Proteins; Public Health; RNA Interference; Regulation; Regulator Genes; Reperfusion Injury; Repression; Research; Role; Sampling; Shapes; Small Interfering RNA; Specificity; Stimulus; Techniques; Therapeutic; Tissues; United States; Validation; Variant; Work; cancer therapy; cardioprotection; chromatin remodeling; cost estimate; epigenetic regulation; epigenomics; experience; experimental study; gene regulatory network; gene repression; heart function; improved; induced pluripotent stem cell derived cardiomyocytes; interest; knock-down; loss of function; mortality; mouse model; novel; patient prognosis; pharmacologic; response; screening; tool; transcription factor; transcriptome sequencing; transcriptomic profiling; transcriptomics; tumor progression; voltage

PROJECT ABSTRACT Epigenetic regulation is critical for cardiac electromechanics and pathology. Epigenetic modulators, such as histone deacetylases (HDACs), are known master regulators of gene expression and influence cardiac function through chromatin remodeling, direct action on transcription factors (TFs), and action on cytoskeletal and contractile proteins, among others. Recently, novel pharmacological agents, HDAC inhibitors, have been developed as treatments for cancer and immune diseases, driving an interest in robust characterization of HDAC control in cardiac function. Our preliminary experiments focused on computational modeling of RNAi-informed transcriptomic data in human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CM) but saw limitations in knockdown efficiency and loss-of-function-only modulation using siRNAs. To extend and improve this work, we propose an experimental approach based on bidirectional perturbation (repression/activation) of individual HDAC genes in hiPSC-CM by CRISPR interference and activation (CRISPRi/a). Transcriptomic analysis of these samples will inform computational gene regulatory network (GRN) inference to model relationships between HDACs, TFs, and cardiac ion channels. GRN-predicted relationships will be validated by all-optical electromechanical assays measuring voltage, calcium, and contraction traces in hiPSC-CM. An iterative approach will allow feedback from functional experiments to refine our computational models. Such studies will advance our understanding of how certain HDACs drive electrophysiological phenotypes in the heart, which is critical in the fields of cardiac injury, cardiac therapeutics, and cardio-oncology.

项目摘要 表观遗传调控对心脏电力学和病理学至关重要。表观遗传调节剂，如 组蛋白脱乙酰酶（HDAC）是已知基因表达的主要调节因子，影响心脏功能 通过染色质重塑，直接作用于转录因子（TF），作用于细胞骨架和 收缩蛋白等。最近，新的药理学试剂，HDAC抑制剂，已经被研究。 作为癌症和免疫性疾病的治疗方法开发，推动了对HDAC的强大表征的兴趣 控制心脏功能。我们的初步实验集中在RNAi信息的计算建模上。 在人诱导多能干细胞衍生的心肌细胞（hiPSC-CM）中的转录组学数据， 在敲低效率和使用siRNA的仅功能丧失调节方面的局限性。扩大和改善 这项工作，我们提出了一种实验方法的基础上双向扰动（抑制/激活）的 通过CRISPR干扰和激活（CRISPRi/a）在hiPSC-CM中的单个HDAC基因。转录组 对这些样本的分析将为计算基因调控网络（GRN）推断提供信息，以建立模型 HDAC、TF和心脏离子通道之间的关系。GRN预测的关系将通过 测量hiPSC-CM中的电压、钙和收缩痕迹的全光机电测定。一个 迭代方法将允许来自功能实验的反馈来改进我们的计算模型。等 研究将促进我们对某些HDAC如何驱动心脏中的电生理表型的理解， 这在心脏损伤、心脏治疗学和心脏肿瘤学领域中是关键的。