Interaction of Galphaq and Kir3, G protein-coupled inwardly rectifying potassium channels.
Activation of substance P receptors, which are coupled to Galpha(q), inhibits the Kir3.1/3.2 channels, resulting in neuronal excitation. We have shown previously that this channel inactivation is not caused by reduction of the phosphatidylinositol 4,5-bisphosphate level in membrane. Moreover, Galpha(q) immunoprecipitates with Kir3.2 (J Physiol 564:489-500, 2005), suggesting that Galpha(q) interacts with Kir3.2. Positive immunoprecipitation, however, does not necessarily indicate direct interaction between the two proteins. Here, the glutathione transferase pull-down assay was used to investigate interaction between Galpha(q) and the K(+) channels. We found that Galpha(q) interacted with N termini of Kir3.1, Kir3.2, and Kir3.4. However, Galpha(q) did not interact with the C terminus of any Kir3 or with the C or N terminus of Kir2.1. TRPC6 is regulated by the signal initiated by Galpha(q). Immunoprecipitation, however, showed that Galpha(q) did not interact with TRPC6. Thus, the interaction between Galpha(q) and the Kir3 N terminus is quite specific. This interaction occurred in the presence of GDP or GDP-AlF(-)(4). The Galpha(q) binding could take place somewhere between residues 51 to 90 of Kir3.2; perhaps the segment between 81 to 90 residues is crucial. Gbetagamma, which is known to bind to N terminus of Kir3, did not compete with Galpha(q) for the binding, suggesting that these two binding regions are different. These findings agree with the hypothesis (J Physiol 564:489-500, 2005) that the signal to inactivate the Kir3 channel could be mainly transmitted directly from Galpha(q) to Kir3.
Kawano
Takeharu
T
Departments of Anatomy & Cell Biology, College of Medicine, University of Illinois at Chicago, Illinois 60612, USA.
Zhao
Peng
P
Floreani
Christina V
CV
Nakajima
Yasuko
Y
Kozasa
Tohru
T
Nakajima
Shigehiro
S
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D050052:TRPC Cation Channels / Q000378:metabolism; D014162:Transfection
D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; C489461:Kir2.1 channel; D024661:Potassium Channels, Inwardly Rectifying; D050052:TRPC Cation Channels; C492145:Trpc6 protein, rat; D043802:GTP-Binding Protein alpha Subunits, Gq-G11