Interaction of Gα_qand Kir3, G Protein-Coupled Inwardly Rectifying Potassium Channels

Abstract

Activation of substance P receptors, which are coupled to G␣ 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, G␣ q immunoprecipitates with Kir3.2 (J Physiol 564: 489 -500, 2005), suggesting that G␣ 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 G␣ q and the K ϩ channels. We found that G␣ q interacted with N termini of Kir3.1, Kir3.2, and Kir3.4. However, G␣ 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 G␣ q . Immunoprecipitation, however, showed that G␣ q did not interact with TRPC6. Thus, the interaction between G␣ q and the Kir3 N terminus is quite specific. This interaction occurred in the presence of GDP or GDP-AlF 4 Ϫ . The G␣ q binding could take place somewhere between residues 51 to 90 of Kir3.2; perhaps the segment between 81 to 90 residues is crucial. G␤␥, which is known to bind to N terminus of Kir3, did not compete with G␣ 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 G␣ q to Kir3.

Specificity of Gβγ Signaling to Kir3 Channels Depends on the Helical Domain of Pertussis Toxin-sensitive Gα Subunits

“…Thus, it is possible that any G␣ subunit can bind to the Kir3 N terminus and sequester G␤␥ from interacting with Kir3, thereby regulating agonist-independent interactions with Kir3. This is consistent with studies showing that both G␣ i and G␣ q can bind to the N terminus of Kir3, whereas only G␣ i exhibited significant binding to the C terminus (51,52,55). Taken together, work from various laboratories described above points to the C terminus of Kir3 as the region that mediates agonist-induced current activation.…”

“…5B were obtained as described under "Experimental Procedures." The G␣ i and G␣ q binding results with the C terminus as well as studies showing that G␣ q can interact with the N but much less with the C terminus of Kir3 (52,55) are all consistent with the interpretation that it is the Kir3 C terminus that is sensitive to the G␣ subunit type. To determine whether the difference in Kir3 C terminus binding to G␣ i and G␣ q can be attributed to the helical domain, we carried out the GST pulldown assay with G␣ qiq and G␣ iqi .…”

“…G␣ subunits were shown to co-immunoprecipitate with the full-length Kir3 channel (51,52). 5 Moreover, G␣ subunits interact directly with Kir3 cytosolic domain fragments in vitro (36,(52)(53)(54)(55). Although both G␣ i and G␣ q can bind the N-terminal domain of Kir3, G␣ i , but not G␣ q or G␣ s , can bind the C terminus of Kir3 (52,55).…”

Gastrin‐releasing peptide acts via postsynaptic BB2 receptors to modulate inward rectifier K+ and TRPV1‐like conductances in rat paraventricular thalamic neurons

“…In theory, all three Kir channel types could be modulated by GRP through BB 2 receptor‐mediated activation of PLC; PLC cleaves the membrane phosphoinositide phosphatidylinositol 4,5‐biphosphate, which is required for their normal physiological properties (Stanfield et al 2002; Jensen et al 2008; Hibino et al 2010). Kir3 channels could also be modulated through PLC‐mediated activation of protein kinase C (Stevens et al 1999), or by a direct interaction of G q /G 11 protein α subunits with the channel (Kawano et al 2007). Kir2 and Kir6, but not Kir3, subfamily members have been detected in PVT (Horio et al 1996; Thomzig et al 2005; Saenz del Burgo et al 2008).…”

Specificity of Gβγ Signaling Depends on Gα Subunit Coupling with G-Protein-Sensitive K⁺ Channels

“…An interesting hypothesis has been put forward to explain the role of G ␣ subunits in controlling G protein signaling [42,43] . This hypothesis suggests that G ␣ i [42] and G ␣ q [44] bind and directly inhibit the activity of Kir3 functions. This G ␣ subunit-mediated inhibition is viewed as being independent of the G ␤ ␥ subunit activation of Kir3.…”