Protein Kinase Novel 2 (PKN2) in heart

心脏中的蛋白激酶 Novel 2 (PKN2)

基本信息

批准号：
10322445
负责人：
Ju Chen
金额：
$ 54.7万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-01-01 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10322445
关键词：
Acute Address Adult Age-Months Angiotensin II Atrial Fibrillation Biological Process Cardiac Cardiac Myocytes Cardiac development Cell physiology Cells Chronic Coronary Arteriosclerosis Defect Development Dilated Cardiomyopathy Disease Embryo Family Heart Heart Abnormalities Heart Diseases Heart Hypertrophy Heart failure Human Immune Knock-out Knockout Mice Left Lymphoid Molecular Morphology Mus Myelogenous Myocardial Infarction Neonatal Patients Phenotype Phosphotransferases Physiological Play Protein Kinase Protein Kinase C Proteins Publishing Quantitative Trait Loci Reporting Risk Role Shapes Signal Transduction Single Nucleotide Polymorphism Skeletal Muscle Smooth Muscle Structure Tamoxifen Therapeutic Ventricular Weaning analog chemical genetics genetic approach genome wide association study heart function inhibitor member mouse model mutant novel phosphoproteomics postnatal postnatal development pressure protective effect ranpirnase small molecule inhibitor therapeutic target vector

项目摘要

PROJECT SUMMARY The Protein Kinase Novel (PKNs) family of kinases, also known as Protein Kinase C-related kinases, belong to the PKC superfamily. A single nucleotide polymorphism at the Pkn2 locus is associated with greater risk for coronary artery disease/myocardial infarction, and elevated levels of PKN2 protein are associated with heart disease, highlighting the importance of PKN2 in heart. Global deletion of Pkn2 in mouse results in lethality at embryonic day (E) 10 with cardiac defects. Conditional deletion of Pkn2, utilizing SM22α-Cre mice, results in partial lethality between E13.5 and weaning, with surviving mutants displaying abnormal cardiac phenotypes. These observations strongly suggest that PKN2 plays a critical role in the developing heart, however, because SM22α-Cre is expressed not only in early developing cardiomyocytes, but also in smooth muscle, skeletal muscle, and myeloid and lymphoid immune cells, this previous study does not address the cardiomyocyte- specific requirement. Intriguingly, a recent study showed that deletion of Pkn1 and Pkn2 in adult cardiomyocytes, utilizing αMHC-MerCreMer mice, did not affect basal cardiac function, but protected mice from pressure overload- and angiotensin II-induced cardiac hypertrophy and heart failure, suggesting that PKN inactivation could be a unique therapeutic target for heart failure. The contradiction between the partial embryonic lethality of SM22α-Cre:Pkn2 knockout mice and protective effects observed in adult αMHC- MerCreMer:Pkn1/2 double knockout mice highlights a critical need to define potential roles of PKN2 in cardiomyocytes at different developmental stages. To address this contradiction, we have generated novel Pkn2 cardiomyocyte-specific constitutive knockout (cKO) and Pkn2 tamoxifen-inducible cardiomyocyte-specific knockout (icKO) mouse models utilizing Xmlc2-Cre and Tnnt2-MerCreMer mouse lines, respectively. Upon preliminary characterization, Pkn2 cKO mice displayed partial postnatal lethality and cardiac morphological defects as early as E12.5. Echocardiographic studies of surviving mutants revealed a dilated cardiomyopathy phenotype in Pkn2 cKO mutants at both 1 and 3 months of age. In contrast to published reports that loss of Pkn2 in adult cardiomyocytes does not affect basal cardiac function, our preliminary observations suggest that Pkn2 deficiency in developing cardiomyocytes is detrimental. Taken together, the above evidence leads us to the hypothesis that PKN2 plays distinct roles at different stages of cardiomyocyte development through the phosphoregulation of specific substrates. Accordingly, our specific aims are: 1. To elucidate the role of PKN2 in cardiomyocytes by analysis of cardiac and cardiomyocyte structure and function in Pkn2 cKO mice, and to identify endogenous substrates of PKN2 in cardiomyocytes by utilizing unbiased phosphoproteomics and a chemical-genetics approach with an analog-sensitive PKN2 mutant, and 2. To determine the cardiomyocyte- specific requirement for PKN2 in postnatal development by analysis of Pkn2 icKO mice.

项目概要蛋白激酶新型 (PKN) 激酶家族，也称为蛋白激酶 C 相关激酶，属于 Pkn2 基因座的单核苷酸多态性与 PKC 超家族的更大风险相关。冠状动脉疾病/心肌梗塞，PKN2 蛋白水平升高与心脏相关疾病，强调了 PKN2 在心脏中的重要性。小鼠中 Pkn2 的整体缺失会导致死亡。使用 SM22α-Cre 小鼠，在胚胎第 10 天（E）有条件地删除 Pkn2，导致出现心脏缺陷。 E13.5 和断奶之间部分致死，存活的突变体表现出异常的心脏表型。然而，这些观察结果强烈表明 PKN2 在发育中的心脏中发挥着关键作用，因为 SM22α-Cre不仅在早期发育的心肌细胞中表达，还在平滑肌、骨骼肌细胞中表达。肌肉、骨髓和淋巴免疫细胞，这项先前的研究并未涉及心肌细胞- 有趣的是，最近的一项研究表明，成人中 Pkn1 和 Pkn2 的缺失。使用 αMHC-MerCreMer 小鼠的心肌细胞不会影响心脏基础功能，但可以保护小鼠免受压力超负荷和血管紧张素 II 诱导的心脏肥大和心力衰竭，表明 PKN 失活可能是心力衰竭的独特治疗靶点。 SM22α-Cre:Pkn2 敲除小鼠的胚胎致死率和在成年 αMHC- 中观察到的保护作用 MerCreMer：Pkn1/2 双敲除小鼠强调了确定 PKN2 的潜在作用的迫切需要为了解决这个矛盾，我们产生了新的心肌细胞。 Pkn2 心肌细胞特异性组成型敲除 (cKO) 和 Pkn2 他莫昔芬诱导型心肌细胞特异性之上初步表征，Pkn2 cKO 小鼠表现出部分产后致死率和心脏形态学早在 E12.5 时，对存活突变体的超声心动图研究就发现了扩张型心肌病。与已发表的报告相比，Pkn2 cKO 突变体在 1 个月和 3 个月时的表型丧失。成人心肌细胞中的 Pkn2 不影响基础心功能，我们的初步观察表明综上所述，上述证据使我们得出结论：心肌细胞发育中的 Pkn2 缺乏是令人痛苦的。 PKN2 在心肌细胞发育的不同阶段发挥不同作用的假设因此，我们的具体目标是： 1. 阐明 PKN2 在磷酸化中的作用。通过分析 Pkn2 cKO 小鼠的心脏和心肌细胞的结构和功能，并利用无偏磷酸蛋白质组学和使用模拟敏感 PKN2 突变体的化学遗传学方法，以及 2. 确定心肌细胞通过对 Pkn2 icKO 小鼠的分析，了解出生后发育中 PKN2 的具体要求。