Endothelial nitric oxide synthase (eNOS) is a key enzyme involved in the regulation of vascular tone and altered eNOS activity leads to endothelial dysfunction and atherosclerosis. Post-transcriptional modulation of eNOS mRNA stability is a major determinant of eNOS expression in vascular endothelium. The molecular mechanisms regulating eNOS mRNA stability remain poorly understood and represent an important gap in knowledge. This competitive renewal application proposes to investigate the molecular mechanisms by which eNOS mRNA stability is regulated. Preliminary studies suggest that the binding of Polypyrimidine Tract Binding Protein 1 (PTB1) to the eNOS mRNA 3?-UTR leads to decreased eNOS mRNA stability and expression. Moreover, we found that PTB1 specifically interacts with the mammalian Ste20-like kinase 1 (Mst1), an upstream serine/threonine-specific protein kinase of the Hippo pathway, which leads to PTB1 phosphorylation and decreased eNOS mRNA stability and protein expression in vascular endothelial cells. Our preliminary data further demonstrate that Mst1 kinase is activated in the atherosclerotic lesions and blocking Mst1 activation by overexpression of dominant negative protein (DN-Mst1) (K59R) in vascular endothelial cells markedly increased eNOS mRNA stability and expression, implicating a functional significance of Mst1 kinase in regulating endothelial functions. We therefore hypothesize that hyperlipidemia-induced activation of Mst1 promotes the binding of PTB1 to eNOS 3?-UTR, which in turn downregulates eNOS expression and causes endothelial dysfunction in the atherogenesis. Accordingly, we propose three comprehensive specific aims to delineate how the regulation of PTB1 by Mst1 kinase affects eNOS mRNA stability and expression, and the extent to which this contributes to the pathogenesis of endothelial dysfunction and vascular disease.
Specific aim 1 will examine the functional significance of Mst1 in regulating eNOS mRNA stability by PTB1. We will identify the Mst1 specific phosphorylation site(s) on PTB1 and then determine whether phosphorylation of PTB1 by Mst1 alters the binding affinity of PTB1 to the eNOS 3?-UTR, leading to decreased eNOS mRNA stability and protein expression in vascular endothelial cells.
Specific aim 2 will determine whether PTB1 induces eNOS mRNA instability by inhibiting 3?-polyadenylation of eNOS mRNA and/or by increasing the binding of microRNAs to the eNOS 3?-UTR.
Specific aim 3 will determine whether the Mst1/PTB1 pathway regulates endothelial dysfunction in vivo. By using our newly generated endothelial specific DN-Mst1 transgenic and Mst1 knockout mice, we will investigate whether eNOS expression, endothelium-dependent vascular relaxation, and atherosclerosis are altered by specifically blocking Mst1 activation in vascular endothelial cells. We envision these will establish the importance of Mst1 as a therapeutic target for improving endothelial function and decreasing cardiovascular disease.
Endothelial dysfunction is a major problem associated with many human diseases, including atherosclerosis, hypertension, heart failure, diabetes, and chronic renal failure, etc. Our studies aim to understand the key factors that cause this problem. Understanding these processes will allow development of new therapies to treat the problem in the future.
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|Yang, Ping; Wei, Xin; Zhang, Jian et al. (2016) Antithrombotic Effects of Nur77 and Nor1 Are Mediated Through Upregulating Thrombomodulin Expression in Endothelial Cells. Arterioscler Thromb Vasc Biol 36:361-9|
|Yi, Bing; Ozerova, Maria; Zhang, Guan-Xin et al. (2015) Post-Transcriptional Regulation of Endothelial Nitric Oxide Synthase Expression by Polypyrimidine Tract-Binding Protein 1. Arterioscler Thromb Vasc Biol 35:2153-60|
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|Qi, Jia; Yang, Ping; Yi, Bing et al. (2015) Heat shock protein 90 inhibition by 17-DMAG attenuates abdominal aortic aneurysm formation in mice. Am J Physiol Heart Circ Physiol 308:H841-52|
|Liu, Yan; Zhang, Jian; Yi, Bing et al. (2014) Nur77 suppresses pulmonary artery smooth muscle cell proliferation through inhibition of the STAT3/Pim-1/NFAT pathway. Am J Respir Cell Mol Biol 50:379-88|
|Qin, Qing; Chen, Ming; Yi, Bing et al. (2014) Orphan nuclear receptor Nur77 is a novel negative regulator of endothelin-1 expression in vascular endothelial cells. J Mol Cell Cardiol 77:20-8|
|Chen, Ming; Yi, Bing; Sun, Jianxin (2014) Inhibition of cardiomyocyte hypertrophy by protein arginine methyltransferase 5. J Biol Chem 289:24325-35|
|Huo, Yan; Yi, Bing; Chen, Ming et al. (2014) Induction of Nur77 by hyperoside inhibits vascular smooth muscle cell proliferation and neointimal formation. Biochem Pharmacol 92:590-8|
|Li, Pan; Liu, Yan; Yi, Bing et al. (2013) MicroRNA-638 is highly expressed in human vascular smooth muscle cells and inhibits PDGF-BB-induced cell proliferation and migration through targeting orphan nuclear receptor NOR1. Cardiovasc Res 99:185-93|
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