Pin1 prolyl isomerase regulates endothelial nitric oxide synthase

描述（由申请人提供）：内皮源性一氧化氮（NO）调节心血管稳态的许多重要方面，血管NO产生的变化少至两倍可对正常血管生理学和血管病理生理学产生重大影响。NO在调节血管舒张状态中具有关键作用，因此在调节血压中具有关键作用。从内皮释放的NO还调节其他过程，包括血小板聚集、血小板和白细胞粘附到内皮、血管平滑肌细胞增殖、内皮细胞凋亡、血管生成和涉及急性炎症的血管渗漏。由于NO在这些过程中的每一个中的关键作用，血管NO产生的异常被认为有助于某些血管疾病如动脉粥样硬化、糖尿病和高血压的发病机制。NO在内皮细胞中通过由内皮型一氧化氮合酶（eNOS）催化的精氨酸氧化而合成。因此，详细了解eNOS酶的结构、功能和调节显然是科学研究的一个有价值的目标。eNOS的表达受两种主要机制的调节：蛋白质-蛋白质相互作用和磷酸化。目前还不清楚这两种不同的eNOS调节过程是如何相互关联的。然而，一段时间以来已知的是，在eNOS中存在丝氨酸116（S116）处的抑制性磷酸化位点。该位点在基础条件下和暴露于促炎细胞因子后在内皮细胞中磷酸化。我们的初步数据产生的假设，即脯氨酸指导的eNOS在S116磷酸化促进结合的Pin 1脯氨酰异构酶和Pin 1诱导的构象变化，抑制eNOS的活性。初步数据还表明，这一过程可能对动脉粥样硬化疾病做出重要贡献。因此，我们建议使用纯化的蛋白质，培养的内皮细胞，完整的血管段，和整个动物，包括动物疾病模型，通过各种不同的方法来测试这些假设。这些研究将提供一个更完整的了解比目前存在的内皮细胞中eNOS的调节，一个非常感兴趣的基础科学家和临床医生的主题。

DESCRIPTION (provided by applicant): Endothelium-derived nitric oxide (NO) regulates many important aspects of cardiovascular homeostasis and alterations in vascular NO production of as little as two-fold can have major consequences in both normal vascular physiology and vascular pathophysiology. NO has a crucial role in regulation of the state of vasodilatation of blood vessels and hence in regulation of blood pressure. NO released from the endothelium also modulates other processes as well including platelet aggregation, platelet and leukocyte adhesion to the endothelium, vascular smooth muscle cell proliferation, endothelial cell apoptosis, angiogenesis, and vascular leakage involved in acute inflammation. Because of the key role of NO in each of these processes, abnormalities in vascular NO production are thought to contribute to the pathogenesis of certain vascular disorders such as those of atherosclerosis, diabetes, and hypertension. NO is synthesized in endothelial cells by oxidation of arginine catalyzed by the enzyme, endothelial nitric oxide synthase (eNOS). Obtaining a detailed understanding of the structure, function, and regulation of eNOS enzyme is thus clearly a worthy goal of scientific investigation. eNOS is regulated post-translationally by two primary mechanisms: protein-protein interactions and phosphorylation. Nothing is yet known about how these two different eNOS regulation processes are interrelated. What has been known for some time, however, is that there exists in eNOS an inhibitory phosphorylation site at serine 116 (S116). This site is phosphorylated in endothelial cells under basal conditions and after exposure to pro-inflammatory cytokines. Our preliminary data give rise to the hypothesis that proline-directed phosphorylation of eNOS at S116 promotes binding of the Pin1 prolyl isomerase and Pin1-induced conformational changes that suppress eNOS activity. Preliminary data also show that this process may make an important contribution to the disease of atherosclerosis. We therefore propose to test these hypotheses by a variety of different methods using purified proteins, cultured endothelial cells, intact blood vessel segments, and whole animals including animal disease models. These studies will provide a more complete understanding than currently exists of regulation of eNOS in endothelial cells, a topic of great interest to both basic scientists and clinicians.