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Functional dissection of the regulatory network that governs cardiomyocyte maturation

控制心肌细胞成熟的调节网络的功能剖析

基本信息

批准号：
10629491
负责人：
Nathan James VanDusen
金额：
$ 24.9万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
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：
2022-09-01 至 2025-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10629491
关键词：
Adult Birth CRISPR screen Cardiac Cardiac Myocytes Cardiomyopathies Cells Complex Congenital Heart Defects Data Defect Deposition Development Disease Disease model Dissection Down-Regulation Epigenetic Process Gene Expression Gene Expression Regulation Genes Genetic Genetic Screening Genetic Transcription Goals Heart Heart failure Histone H2B Homeostasis Human Hypertrophic Cardiomyopathy Hypertrophy Incidence Maps Mentors Metabolism Molecular Morphology Mus Mutate Mutation Myosin ATPase Myosin Heavy Chains Neonatal Operative Surgical Procedures Palliative Surgery Pathologic Patients Phase Phenotype Process Production Protein Isoforms Protocols documentation Regenerative Medicine Regulation Regulatory Element Replacement Therapy Repression Role Sarcomeres Stress Ubiquitination chromatin modification conditional knockout congenital heart disorder experimental study heart function hemodynamics histone modification improved in vivo insight loss of function molecular targeted therapies neonate novel promoter regenerative approach response sound targeted treatment tool transcription regulatory network

项目摘要

ABSTRACT Between birth and adulthood, cardiomyocytes (CMs) undergo profound changes in size, ultrastructure, metabolism, and gene expression, a process collectively referred to as CM maturation. Although highly coordinated, the transcriptional network that governs this process is not understood. This lack of understanding is a barrier to cardiac regenerative medicine, where our current inability to mature CMs differentiated from non- myocytes limits their use for disease modeling or replacement therapy. In addition, disruption of maturation by abnormal hemodynamic loads in neonates who have undergone surgery to correct congenital heart defects likely contributes to their high incidence of heart failure in adulthood. A sound understanding of the regulatory network governing CM maturation will inspire hypothesis driven attempts to surmount these challenges. In mice, a key hallmark of CM maturation is sarcomere isoform switching, including the well documented neonatal switch from Myosin Heavy Chain 7 (Myh7) to Myosin Heavy Chain 6 (Myh6). We have conducted and validated an in vivo high throughput CRISPR screen for transcriptional regulators of CM maturation, using the Myh7/6 isoform switch as the readout. Two top candidates from this screen, Rnf20 and Rnf40, form a complex which governs deposition of the poorly understood chromatin modification histone-2B mono-ubiquitination (H2Bub1). H2Bub1 function is unexplored in the heart, and associated with congenital heart disease mutations in human patients. Therefore, In Aim 1 we will perform a detailed characterization of the impact of Rnf20/40 on genetic regulation of CM maturation. A major reason why the transcriptional networks controlling CM maturation are poorly understood is that appropriate tools were previously unavailable. In this proposal we will utilize a suite of novel in vivo tools to begin mapping the transcriptional regulatory networks that govern CM maturation. In Aim 2 we will seek to accomplish this goal by dissecting two Myh7 cis-regulatory elements. Furthermore, we will contrast the genetic regulation of CM maturation with pathological hypertrophy by discovering and describing the key regulatory mechanisms of Myh7, which is deactivated during maturation and re-activated in response to pathological stress. These experiments will yield mechanistic clues as to how gene expression is coordinately controlled during maturation and disease, thus enabling improved CM production protocols and targeted therapies. Collectively these experiments will begin to map the transcriptional regulatory network that governs maturation by defining the functions of select trans-acting regulators and cis-regulatory elements.

摘要