Regulation of Cardiac Remodeling by SRF-mAKAP complexes

SRF-mAKAP 复合物对心脏重构的调节

基本信息

批准号：
8866455
负责人：
CATHERINE LAURA PASSARIELLO
金额：
$ 4.35万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-07-15 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8866455
关键词：
A kinase anchoring protein Adrenergic Agents Adult Affect Affinity Angiotensins Binding Calcium Cardiac Cardiac Myocytes Cell Nucleus Cells Cessation of life Complex Contractile Proteins Coupled Cytoskeletal Proteins Cytosol DNA Binding Data Development Diagnosis Enzymes Exhibits Extracellular Signal Regulated Kinases Family member Gene Expression Genes Genetic Genetic Transcription Growth Health Heart Heart Diseases Heart Hypertrophy Heart failure Hypertrophy In Vitro Infusion procedures Knockout Mice Length MAPK3 gene MEKKs MEKs Maps Mitogen-Activated Protein Kinases Modeling Mus Muscle Muscle Cells Neonatal Neurosecretory Systems Nuclear Envelope PDPK1 gene Pathway interactions Peptides Phenotype Phenylephrine Phosphorylation Physiological Prevention Protein Binding Pump RNA Interference RPS6KA gene Rattus Regimen Regulation Research Infrastructure Ribosomal Protein S6 Kinase Risk Risk Factors Scaffolding Protein Serum Response Factor Signal Pathway Signal Transduction Signaling Molecule Specificity Stress Syndrome Tamoxifen Testing Therapeutic Ventricular adrenergic constriction cytokine design domain mapping in vivo insight mortality muscle stress mutant novel novel therapeutic intervention pressure protein complex protein expression receptor recombinase research study response scaffold transcription factor

项目摘要

DESCRIPTION (provided by applicant): Cardiac hypertrophy is the main compensatory response to stress in heart disease and a leading risk for the development of heart failure. In the adult heart, the serum response factor (SRF) transcription factor is an important regulator of myocyte gene expression, directing the transcription of genes encoding cytoskeletal and contractile proteins, calcium-handling molecules and factors controlling myocyte phenotype. In this application, I examine a novel mechanism for the control of the ER-RSK3-SRF pathway by the protein scaffold muscle A-kinase anchoring protein � (mAKAP�). The Kapiloff lab has shown that mAKAP� is required in vitro for adrenergic and cytokine-induced neonatal myocyte hypertrophy. Preliminary Data suggest that mAKAP� anchored RSK3 is important for the phosphorylation of SRF S103 and induction of myocyte hypertrophy. In this application, I test the hypothesis that hypertrophic gene expression regulated by SRF is dependent upon SRF association with mAKAP� complexes. I propose to examine how SRF binding to mAKAP� facilitates RSK3-catalyzed SRF phosphorylation and activity and to test in vivo whether the mAKAP� scaffold is in fact required for concentric cardiac hypertrophy. Specific Aim 1: mAKAP�-SRF Signalosomes. mAKAP� and SRF can be co-immunoprecipitated using adult heart extracts. I propose to map the domain on mAKAP� that binds SRF to determine whether their interaction is direct. The mapping experiments are designed to identify a small peptide that can compete endogenous mAKAP�-SRF binding in cells, as well as to allow the construction of a full-length mAKAP� SRF-binding mutant In addition, using RNA interference and a currently available peptide that competes RSK3 binding to mAKAP�, I will test whether mAKAP� expression and mAKAP�-RSK3 binding is important for �-adrenergic- induced SRF DNA binding and transcriptional activity. Specific Aim 2: The requirement for mAKAP� in pathological remodeling in vivo. I propose to stress a new cardiac-specific mAKAP� knock-out mouse by transverse aortic constriction and phenylephrine/angiotensin infusion via osmotic pump. I predict that in response to both pressure overload and neuroendocrine over-stimulation that the mAKAP� knock-out mouse will exhibit less concentric cardiac myocyte hypertrophy and decreased overall cardiac remodeling. I also predict that SRF transcriptional activity will be reduced upon genetic deletion The first Specific Aim will provide novel, detailed mechanistic insights into the proposed ERK-RSK3- SRF pathway, including the specific association of these molecules with mAKAP�. The second Specific Aim will show the physiologic relevance of the related scaffold protein. Together, my experiments should reveal a novel therapeutic approach for the prevention or treatment of heart failure, the targeting of critical scaffold proteins that provide the architectral infrastructure for the hypertrophic signaling network.

描述（由应用提供）：心脏肥大是对心脏病压力的主要补偿性反应，也是发展心力衰竭的领先风险。在成人心脏，血清反应因子（SRF）转录因子是肌细胞基因表达的重要调节剂，指导编码细胞骨架和收缩蛋白的基因转录，钙处理分子和控制心肌细胞表型的因子。在此应用中，我检查了一种新的机制，可以通过蛋白质支架肌肉A-激酶锚定蛋白（makap）控制ER-RSK3-SRF途径。 Kapiloff实验室表明，肾上腺因子和细胞因子诱导的新生儿肌细胞肥大需要MAKAP。初步数据表明，MAKAP锚定RSK3对于SRF S103的磷酸化和肌细胞肥大的诱导很重要。在此应用中，我检验了以下假设：由SRF调节的肥厚基因表达取决于SRF与MAKAP的关联是复杂的。我建议检查SRF与MAKAP的结合如何促进RSK3催化的SRF磷酸化和活性，并在体内测试MAKAP支架是否实际上是同心心脏肥大所需的。特定目标1：MAKAP信号体。 MAKAP和SRF可以使用成人心脏提取物共免疫沉淀。我建议在makap上绘制绑定SRF的域以确定其相互作用是否直接。 The mapping experiments are designed to identify a small pepper that can compete endogenous mAKAP‐SRF binding in cells, as well as to allow the construction of a full-length mAKAP‐ SRF binding mutant In addition, using RNA interference and a currently available peptide that competes RSK3 binding to mAKAP‐, I will test whether mAKAP‐ expression and mAKAP‐RSK3 binding is important for ‐adrenergic- induced SRF DNA结合和转录活性。特定目的2：在体内病理重塑中MAKAP的要求。我建议通过横向主动脉收缩和通过渗透泵输注弯曲的小鼠敲除小鼠中的新心脏特异性MAKAP。我预测，响应压力超负荷和神经内分泌过度刺激，MAKAP会杀死浓缩的心肌肥大和改善的整体心脏重塑。我还预测，在遗传缺失时SRF转录活性将减少，第一个特定目标将对所提出的ERK-RSK3-SRF途径提供新颖的，详细的机械见解，包括这些分子与MAKAP的特定关联。第二个特定目的将显示相关支架蛋白的生理相关性。总之，我的实验应该揭示一种用于预防或治疗心力衰竭的新型治疗方法，即关键支架蛋白的靶向，该临界脚手蛋白为肥厚信号网络提供建筑基础设施。