Identification of an integration center for cross-talk between protein kinase C and G protein modulation of N-type calcium channels.
The modulation of presynaptic calcium channel activity by second messengers provides a fine tuning mechanism for neurotransmitter release. In neurons, the activation of certain G protein-coupled receptors reduces N-type channel activity by approximately 60%. In contrast, activation of protein kinase C (PKC) results in an approximately 50% increase in N-type channel activity, and subsequent G protein inhibition is antagonized. Here, we describe the molecular determinants that control the dual effects of PKC-dependent phosphorylation. The double substitution of two adjacent PKC consensus sites in the calcium channel domain I-II linker (Thr422, Ser425) to alanines abolished both PKC-dependent up-regulation and the PKC-G protein cross-talk. The single substitution of Ser425 to glutamic acid abolished PKC up-regulation but had no effect on G protein modulation. Replacement of Thr422 with glutamic acid eliminated PKC-dependent up-regulation and mimicked the effects of PKC phosphorylation on G protein inhibition. Our data suggest that Thr422 mediates the antagonistic effect of PKC on G protein modulation, while phosphorylation of either Thr422 or Ser425 are sufficient to increase N-type channel activity. Thus, Thr422 serves as a molecular switch by which PKC is able to simultaneously trigger the up-regulation of channel activity and antagonize G protein inhibition.
Department of Pharmacology and Therapeutics, Neuroscience Research Group, University of Calgary, Calgary, Alberta T2N 4N1, Canada.
J
J
Hamid
D
D
Nelson
R
R
Spaetgens
S J
SJ
Dubel
T P
TP
Snutch
G W
GW
Zamponi
D000595:Amino Acid Sequence; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004594:Electrophysiology; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009474:Neurons / Q000502:physiology*; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D015398:Signal Transduction; D009435:Synaptic Transmission / Q000502:physiology*; D014162:Transfection
D015220:Calcium Channels; D011493:Protein Kinase C; D019204:GTP-Binding Proteins