Cantrell Angela R, Tibbs Victoria C, Yu Frank H, Murphy Brian J, Sharp Elizabeth M, Qu Yusheng, Catterall William A, Scheuer Todd
Department of Pharmacology, University of Washington, Seattle 98195-7280, USA.
Mol Cell Neurosci. 2002 Sep;21(1):63-80. doi: 10.1006/mcne.2002.1162.
Activation of D1-like dopamine (DA) receptors reduces peak Na(+) current in hippocampal neurons voltage-dependent in a manner via phosphorylation of the alpha subunit. This modulation is dependent upon activation of cAMP-dependent protein kinase (PKA) and requires phosphorylation of serine 573 (S573) in the intracellular loop connecting homologous domains I and II (L(I-II)) by PKA anchored to A kinase anchoring protein-15 (AKAP-15). Activation of protein kinase C (PKC) also reduces peak Na(+) currents and enhances the strength of the PKA modulatory pathway. Here we probe the molecular mechanism responsible for the convergent effects of PKA and PKC on brain Na(v)1.2a channels. Analysis of the interaction of AKAP-15 with the intracellular loops of the Na(v)1.2a channel shows that it binds to L(I-II), thereby targeting PKA directly to its sites of phosphorylation on the Na(+) channel by specific protein-protein interactions. Mutagenesis and expression experiments indicate that reduction of peak Na(+) current by PKC requires S554 and S573 in L(I-II) in addition to S1506 in the inactivation gate. In addition, PKC-dependent phosphorylation of S576 in L(I-II) is necessary for enhancement of PKA modulation of brain Na(+) channels. When S576 is phosphorylated by PKC, the increase in modulation by PKA activation requires phosphorylation of S687 in L(I-II). Thus, the maximal modulation of these Na(+) channels by concurrent activation of PKA and PKC requires phosphorylation at four distinct sites in L(I-II): S554, S573, S576, and S687. This convergent regulation provides a novel mechanism by which information from multiple signaling pathways may be integrated at the cellular level in the hippocampus and throughout the central nervous system.
D1样多巴胺（DA）受体的激活通过α亚基的磷酸化，以一种电压依赖性方式降低海马神经元中的峰值钠电流。这种调节依赖于环磷酸腺苷（cAMP）依赖性蛋白激酶（PKA）的激活，并且需要锚定在A激酶锚定蛋白-15（AKAP-15）上的PKA对连接同源结构域I和II的细胞内环（L（I-II））中的丝氨酸573（S573）进行磷酸化。蛋白激酶C（PKC）的激活也会降低峰值钠电流，并增强PKA调节途径的强度。在此，我们探究负责PKA和PKC对脑Na（v）1.2a通道产生趋同效应的分子机制。对AKAP-15与Na（v）1.2a通道细胞内环相互作用的分析表明，它与L（I-II）结合，从而通过特异性蛋白质-蛋白质相互作用将PKA直接靶向到其在钠通道上的磷酸化位点。诱变和表达实验表明，PKC降低峰值钠电流除了需要失活门中的S1506外，还需要L（I-II）中的S554和S573。此外，L（I-II）中S576的PKC依赖性磷酸化对于增强脑钠通道的PKA调节是必要的。当S576被PKC磷酸化时，PKA激活导致的调节增加需要L（I-II）中的S687磷酸化。因此，PKA和PKC同时激活对这些钠通道的最大调节需要L（I-II）中四个不同位点的磷酸化：S554、S573、S576和S687。这种趋同调节提供了一种新机制，通过该机制来自多个信号通路的信息可以在海马体以及整个中枢神经系统的细胞水平上进行整合。
