title: Cardiac calcium channels expressed in Xenopus oocytes are modulated by dephosphorylation but not by cAMP-dependent phosphorylation.
abstract: Enhancement of cardiac L-type Ca2+ channel activity by norepinephrine via phosphorylation by protein kinase A (PKA) underlines the positive inotropic effect of this transmitter and is a classical example of an ion channel modulation. However, it is not clear whether the channel protein itself (and which subunit) is a substrate for PKA. We have expressed various combinations of the cardiac Ca2+ channel subunits in Xenopus oocytes by injecting subunit mR-NAs. Expression of beta or alpha 2/delta + beta subunits potentiated the native (endogenous) Ca2+ channel currents in the oocyte (similar to T or N but not L-type). This potentiated endogenous current was enhanced by intracellular injection of cAMP or of the catalytic subunit of PKA, and this effect was reversed by the injection of a PKA inhibitor suggesting the presence of basal phosphatase activity. When a cardiac channel of alpha 1 + beta, alpha 1 + alpha 2/delta or alpha 1 + alpha 2/delta + beta composition was expressed at levels high enough that the contribution of the endogenous current became negligible, cAMP and PKA failed to increase the Ca2+ channel current, whereas PKA inhibitors and the catalytic subunit of protein phosphatase 1 reduced the amplitude of the current. Reduction of the current by PKA inhibitors was observed regardless of the presence of the beta subunit, suggesting a major role for the alpha 1 subunit in this process. These results suggest that, like in the heart, when expressed in Xenopus oocytes, the cardiac L-type Ca2+ channels are phosphorylated in basal state and dephosphorylation reduces their activity. However, unlike the situation in the heart, the activity of the channel cannot be enhanced by PKA-catalyzed phosphorylation, suggesting that the channel is already fully phosphorylated in its basal state.
authors.affiliation: Department of Physiology and Pharmacology, Sackler School of Medicine, Tel Aviv University, Ramat Aviv, Israel.
authors.forename: D
authors.initials: D
authors.lastname: Singer-Lahat
authors.forename: I
authors.initials: I
authors.lastname: Lotan
authors.forename: M
authors.initials: M
authors.lastname: Biel
authors.forename: V
authors.initials: V
authors.lastname: Flockerzi
authors.forename: F
authors.initials: F
authors.lastname: Hofmann
authors.forename: N
authors.initials: N
authors.lastname: Dascal
mesh_terms: D000818:Animals; D016477:Artifacts; D001464:Barium / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000008:administration & dosage / Q000037:antagonists & inhibitors / Q000378:metabolism; D018076:DNA, Complementary / Q000008:administration & dosage / Q000235:genetics; D005260:Female; D008845:Microinjections; D008954:Models, Biological; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D009206:Myocardium / Q000737:chemistry*; D009638:Norepinephrine / Q000494:pharmacology; D009865:Oocytes; D009928:Organ Specificity; D010749:Phosphoprotein Phosphatases / Q000008:administration & dosage / Q000378:metabolism; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000008:administration & dosage / Q000235:genetics; D011817:Rabbits; D015854:Up-Regulation; D014982:Xenopus laevis
chemical_list: D015220:Calcium Channels; D018076:DNA, Complementary; D009124:Muscle Proteins; D012333:RNA, Messenger; D001464:Barium; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D002118:Calcium; D009638:Norepinephrine