Norepinephrine Transport Regulation By Phosphorylation

通过磷酸化调节去甲肾上腺素运输

• 批准号: 7347631
• 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/7347631
• 关键词: Affect; Antibodies; Attention; Attenuated; Binding; Biotinylation; Brain Diseases; Cardiovascular Diseases; Cardiovascular system; Catecholamines; Cell Line; Cell model; Cells; Dependency; Development; Down-Regulation; Heart Rate; Human; Ions; Kinetics; Label; Link; Maintenance; Mediating; Membrane; Membrane Microdomains; Metabolic; Molecular; Motivation; Mutation; Norepinephrine; Phenotype; Phosphopeptides; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Physiological Processes; Placenta; Pre-Eclampsia; Pregnancy; Property; Protein Kinase C; Proteins; RNA Interference; Rattus; Receptor Activation; Regulation; Research; Research Personnel; Resistance; Role; Second Messenger Systems; Signal Transduction; Signaling Molecule; Site; Substance P; Substance P Receptor; System; Testing; Time; Tricyclic Antidepressive Agents; base; citrate carrier; coping; extracellular; genetic variant; macromolecule; mutant; noradrenaline transporter; novel therapeutics; prevent; programs; psychostimulant; receptors for activated C kinase; research study; second messenger; syntaxin 1A; trafficking; trophoblast; vasoconstriction

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生理过程。 NE信号通过包括NE转运蛋白（NETS）在内的多种大分子（NETS）动态调节。在中枢神经系统和外围表达的网是三环抗抑郁药和精神刺激物的重要靶标。 NE转运的改变记录在心血管疾病和脑部疾病中。在某些心血管表型中，第二使者链接的激酶介导的NET遗传变异的调节已改变，这意味着寻找NE转运调控的潜在机制。 NET包含了几种激酶（包括蛋白激酶C（PKC））的推定磷酸化位点。最近，我们证明，在大鼠胎盘滋养细胞中，PKC激活通过脂质筏增强了天然净内在化并刺激转运蛋白磷酸化。根据对天然净调控的研究，我们指示我们的研究探索净磷酸化在NE运输调节中的作用。使用表达HNET的人胎盘滋养细胞细胞系（HTR-HNET细胞），我们证明了HNET预测的PKC磷酸化基序的双突变可防止神经激素1受体1受体（NK1R）/PKC介导的介导的转运蛋白调节和磷酸化。过度的神经新闻素分泌与先兆子痫有关，因此，NK1R激活对胎盘净的调节表明这种调节在维持正常妊娠时的生理相关性。基于这些观察结果，我们建议检验一个特定的假设，即在两个独立的特定目标中，NE运输调节需要净磷酸化。在特定目标I中，我们将确定NK1R/PKC涉及的信号，特定位点和基序调节净磷酸化和表达，以探索净磷酸化与NE转运之间的关系。具体目标II将检验以下假设：NK1R/PKC调节的净磷酸化和与转运蛋白相关蛋白的相互作用发生在脂质筏中，并且该关联以活动依赖性的方式建立了转运蛋白的分布和功能，以应对降压信号的需求。这项研究的结果将提供可能用于开发旨在用于治疗心血管疾病和脑部疾病的新型治疗策略的信息。