Garcia D E, Brown S, Hille B, Mackie K
Department of Physiology and Biophysics, University of Washington, Seattle, Washington 98195, USA.
J Neurosci. 1998 Apr 15;18(8):2834-41. doi: 10.1523/JNEUROSCI.18-08-02834.1998.
We have found that phosphorylation of a G-protein-coupled receptor by protein kinase C (PKC) disrupts modulation of ion channels by the receptor. In AtT-20 cells transfected with rat cannabinoid receptor (CB1), the activation of an inwardly rectifying potassium current (Kir current) and depression of P/Q-type calcium channels by cannabinoids were prevented by stimulation of protein kinase C by 100 nM phorbol 12-myristate 13-acetate (PMA). In contrast, activation of Kir current by somatostatin was unaffected, and inhibition of calcium channels was only modestly attenuated. The possibility that PKC acted by phosphorylating CB1 receptors was confirmed by demonstrating that PKC phosphorylated a single serine (S317) of a fusion protein incorporating the third intracellular loop of CB1. Mutating this serine to alanine did not affect the ability of CB1 to modulate currents, but it eliminated disruption by PMA, demonstrating that PKC can disrupt ion channel modulation by receptor phosphorylation.
我们发现，蛋白激酶C（PKC）对G蛋白偶联受体的磷酸化会破坏该受体对离子通道的调节作用。在用大鼠大麻素受体（CB1）转染的AtT-20细胞中，100 nM佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）刺激蛋白激酶C可阻止大麻素对内向整流钾电流（Kir电流）的激活以及对P/Q型钙通道的抑制。相比之下，生长抑素对Kir电流的激活未受影响，钙通道的抑制仅略有减弱。通过证明PKC使包含CB1第三个细胞内环的融合蛋白的单个丝氨酸（S317）磷酸化，证实了PKC通过磷酸化CB1受体发挥作用。将该丝氨酸突变为丙氨酸不影响CB1调节电流的能力，但消除了PMA的破坏作用，表明PKC可通过受体磷酸化破坏离子通道调节。
