AKAP Anchored PKA and Calcineurin Regulation of Neuronal L-type Calcium Channels

AKAP 锚定 PKA 和钙调磷酸酶对神经元 L 型钙通道的调节

• 批准号: 8197228
• 负责人: MARK L DELL'ACQUA
• 金额: $ 33.95万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-12-15 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8197228
• 关键词: A kinase anchoring protein; Address; Adenylate Cyclase; Adrenergic Receptor; Affinity; Aging; Alzheimer' s Disease; Binding; Biochemical; Biological; CREB1 gene; Calcineurin; Calcium Channel; Calcium ion; Calcium/calmodulin-dependent protein kinase; Calmodulin; Cardiac Myocytes; Cell Nucleus; Cell Survival; Cell membrane; Cells; Complex; Coupling; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Dendrites; Dendritic Spines; Development; Distal; Down Syndrome; Enzymes; Equilibrium; Event; Excitatory Synapse; Feedback; Gene Expression; Genetic Transcription; Heart; Hippocampus (Brain); Impaired cognition; L-Type Calcium Channels; L-type calcium channel alpha(1C); Lead; Learning; Leucine Zippers; Mediating; Memory; Mental Retardation; Modeling; Molecular; Muscle Cells; Myocardial Contraction; Nerve Degeneration; Neuronal Plasticity; Neurons; Nuclear; Pathway interactions; Phosphorylation; Phosphotransferases; Physiological; Play; Positioning Attribute; Production; Protein Kinase; Protein phosphatase; Proteins; Recruitment Activity; Regulation; Role; Scaffolding Protein; Signal Transduction; Site; Surface; Synaptic plasticity; System; Testing; calcineurin phosphatase; computerized data processing; molecular assembly/self assembly; neuronal survival; novel; novel therapeutics; postsynaptic; protein complex; reconstitution; response; scaffold; transcription factor; voltage

Elucidating mechanisms regulating neuronal survival and plasticity is relevant for understanding normal learning and memory as well as cognitive impairments in mental retardation, aging and Alzheimer¿s. Influx of calcium ions (Ca2+) through L-type voltage-gated calcium channels (LTCCs) can influence long-term changes in synaptic plasticity and neuronal survival by turning on and off gene transcription in the nucleus. While it is known that signaling very near the site of Ca2+ influx is required for regulation of both LTCC activity and gene expression, molecular mechanisms that organize channel proximal signals and transduce them to the nucleus are largely unknown. One important pathway by which LTCC activity in neurons is regulated involves b-adrenergic receptor-mediated stimulation of cAMP production by adenylyl cyclase and activation of the kinase PKA. Previous studies in the heart suggest that efficient regulation of LTCC activity by PKA requires phosphorylation of the channel protein and localization of PKA near the channel through binding to A-kinase-anchoring proteins (AKAP). However, little is known about the roles of AKAPs or the opposing actions of protein phosphatases in neuronal LTCC regulation. In postsynaptic neurons one AKAP that may play a key role in regulating LTCC phosphorylation and signaling to transcription factors in the nucleus is AKAP79/150. Our overall hypothesis is that AKAP79/150 targets PKA and CaN to LTCCs to bi-directionally regulate channel activity and signaling to the nucleus. We will test this hypothesis in the context of a model in which anchored CaN strongly opposes cAMP-PKA regulation of the channel currents to function as a Ca2+ negative feedback mechanism. In addition, we will explore a novel role for dynamic anchoring of PKA and CaN to AKAP79/150 in these plasma membrane localized Ca2+ signaling events that also control downstream activation of NFAT and CREB transcription factors. Thus, our studies will characterize a novel molecular assembly that coordinates plasma membrane LTCC Ca2+ signaling to regulate both local and distal responses that are important in neuronal plasticity. We will use biochemical, cell biological and electrophysiological approaches in HEK-293 cells and hippocampal neurons to study AKAP79/150-LTCC regulation: (Aim 1) Molecular and functional characterization of a direct interaction between AKAP79/150 and the LTCC CaV1.2 in neuronal channel regulation; (Aim 2) Role of dynamic PKA and CaN anchoring to AKAP79/150 in neuronal LTCC regulation; (Aim 3) Role of the AKAP79/150 channel-associated signaling complex in regulating neuronal LTCC excitation-transcription coupling.

阐明调节神经元存活和可塑性的机制与 了解正常的学习和记忆以及认知障碍， 迟缓、衰老和老年痴呆症。钙离子（Ca 2+）通过L型电压门控 钙通道（LTCCs）可以影响突触可塑性和神经元的长期变化， 通过开启和关闭细胞核中的基因转录来维持生存。虽然已知发信号 非常接近Ca 2+内流的位点是调节LTCC活性和基因表达所必需的。 表达，组织通道近端信号并使其稳定的分子机制 对原子核的作用还不太清楚神经元中LTCC活性的一个重要途径 调节涉及b-肾上腺素能受体介导的cAMP产生的刺激， 腺苷酸环化酶和激酶PKA的活化。先前的心脏研究表明， PKA对LTCC活性的有效调节需要通道蛋白的磷酸化， PKA通过与A-激酶锚定蛋白（AKAP）结合而定位在通道附近。 然而，关于AKAP的作用或蛋白质的相反作用知之甚少 磷酸酶在神经元LTCC调节中的作用。在突触后神经元中，一种AKAP可能起着 在调节LTCC磷酸化和向细胞核中转录因子发出信号中的关键作用 AKAP79/150我们的总体假设是AKAP 79/150靶向PKA和CaN至LTCC， 双向调节通道活性和向细胞核的信号传导。我们将测试这个 锚定CaN强烈反对cAMP-PKA的模型背景下的假设 调节通道电流以起到Ca 2+负反馈机制的作用。在 此外，我们将探索PKA和CaN动态锚定AKAP 79/150的新作用， 这些质膜定位的Ca 2+信号传导事件也控制下游激活， NFAT和CREB转录因子。因此，我们的研究将表征一种新的分子 一种协调质膜LTCC Ca 2+信号传导以调节局部和 在神经元可塑性中很重要的远端反应。我们将使用生物化学，细胞生物学 和电生理方法在HEK-293细胞和海马神经元中研究 AKAP 79/150-LTCC调控：（目的1）直接调控的分子和功能表征 AKAP 79/150和LTCC CaV1.2在神经元通道调节中的相互作用;（目的2） 动态PKA和CaN锚定AKAP 79/150在神经元LTCC调节中的作用;（目的3） AKAP 79/150通道相关信号复合物在神经元LTCC调控中的作用 激发-转录偶联。