Norepinephrine Transport Regulation By Phosphorylation

通过磷酸化调节去甲肾上腺素运输

• 批准号: 7196942
• 负责人: LANKUPALLE D JAYANTHI
• 金额: $ 22.05万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-02-01 至 2012-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7196942
• 关键词: Norepinephrine; Transport; Regulation; Phosphorylation

DESCRIPTION (provided by applicant): The catecholamine, norepinephrine (NE) governs a spectrum of physiologic processes from vasoconstriction and heart rate to attention and motivation. NE signaling is dynamically regulated by a diverse set of macromolecules including NE transporters (NETs). NETs expressed in the CNS and periphery are important targets for tricyclic antidepressants and psychostimulants. Altered NE transport is documented in cardiovascular diseases and brain disorders. Second messenger linked kinase mediated regulation of human genetic variants of NET is altered in some cardiovascular phenotypes signifying the search for underlying mechanisms of NE transport regulation. NET contains putative phosphorylation sites for several kinases including protein kinase C (PKC). Recently, we demonstrated that, in rat placental trophoblasts, PKC activation enhances native NET internalization via lipid rafts and stimulates transporter phosphorylation. Following the studies on native NET regulation, we directed our studies to explore the role of NET phosphorylation in NE transport regulation. Using a human placental trophoblast cell line expressing hNET (HTR-hNET cells), we demonstrate that a double mutation at a predicted PKC phosphorylation motif of hNET prevents neurokinin 1 receptor (NK1R)/PKC-mediated transporter regulation and phosphorylation. Excessive neurokinin secretion is linked to pre-eclampsia, and thus, the regulation of placental NET by NK1R activation suggests physiological relevance of such regulation in the maintenance of a normal pregnancy. Based on these observations, we propose to test a specific hypothesis that NET phosphorylation is required for NE transport regulation in two separate Specific Aims. In Specific Aim I, we will identify the signals, specific sites and motifs involved in NK1R/PKC regulated NET phosphorylation and expression to explore the relationship between NET phosphorylation and NE transport. Specific Aim II will test the hypothesis that NK1R/PKC regulated NET phosphorylation and interaction with transporter-associated proteins occur in lipid rafts, and that this association establishes transporter distribution and function in an activity-dependent manner to cope with the demands in the milieu of incoming signals. The results from this research will provide information that could be of use in the development of new therapeutic strategies aimed at NE transport regulation in the treatment of both cardiovascular diseases and brain disorders.

描述（由申请人提供）：儿茶酚胺、去甲肾上腺素（NE）控制着从血管收缩和心率到注意力和动机的一系列生理过程。 NE 信号传导受到包括 NE 转运蛋白 (NET) 在内的多种大分子的动态调节。中枢神经系统和外周表达的 NET 是三环类抗抑郁药和精神兴奋剂的重要靶点。心血管疾病和脑部疾病中记录了 NE 转运的改变。第二信使相关激酶介导的人类 NET 遗传变异的调节在一些心血管表型中发生改变，这意味着寻找 NE 转运调节的潜在机制。 NET 包含多种激酶的假定磷酸化位点，包括蛋白激酶 C (PKC)。最近，我们证明，在大鼠胎盘滋养层中，PKC 激活可通过脂筏增强天然 NET 内化并刺激转运蛋白磷酸化。在对天然 NET 调节的研究之后，我们的研究方向是探索 NET 磷酸化在 NE 运输调节中的作用。使用表达 hNET 的人胎盘滋养层细胞系（HTR-hNET 细胞），我们证明 hNET 的预测 PKC 磷酸化基序的双突变可阻止神经激肽 1 受体 (NK1R)/PKC 介导的转运蛋白调节和磷酸化。神经激肽分泌过多与先兆子痫有关，因此，NK1R 激活对胎盘 NET 的调节表明这种调节在维持正常妊娠中的生理相关性。基于这些观察，我们建议在两个单独的特定目标中测试一个特定的假设，即 NET 磷酸化是 NE 运输调节所必需的。在Specific Aim I中，我们将鉴定NK1R/PKC调节NET磷酸化和表达所涉及的信号、特定位点和基序，以探讨NET磷酸化和NE转运之间的关系。具体目标 II 将检验以下假设：NK1R/PKC 调节脂筏中发生的 NET 磷酸化和与转运蛋白相关蛋白的相互作用，并且这种关联以活性依赖的方式建立转运蛋白的分布和功能，以应对传入信号环境中的需求。这项研究的结果将提供可用于开发新的治疗策略的信息，该策略旨在治疗心血管疾病和脑部疾病中的 NE 转运调节。