title: N-terminal tyrosine residues within the potassium channel Kir3 modulate GTPase activity of Galphai.
abstract: trkB activation results in tyrosine phosphorylation of N-terminal Kir3 residues, decreasing channel activation. To determine the mechanism of this effect, we reconstituted Kir3, trkB, and the mu opioid receptor in Xenopus oocytes. Activation of trkB by BDNF (brain-derived neurotrophic factor) accelerated Kir3 deactivation following termination of mu opioid receptor signaling. Similarly, overexpression of RGS4, a GTPase-activating protein (GAP), accelerated Kir3 deactivation. Blocking GTPase activity with GTPgammaS also prevented Kir3 deactivation, and the GTPgammaS effect was not reversed by BDNF treatment. These results suggest that BDNF treatment did not reduce Kir3 affinity for Gbetagamma but rather acted to accelerate GTPase activity, like RGS4. Tyrosine phosphatase inhibition by peroxyvanadate pretreatment reversibly mimicked the BDNF/trkB effect, indicating that tyrosine phosphorylation of Kir3 may have caused the GTPase acceleration. Tyrosine to phenylalanine substitution in the N-terminal domain of Kir3.4 blocked the BDNF effect, supporting the hypothesis that phosphorylation of these tyrosines was responsible. Like other GAPs, Kir3.4 contains a tyrosine-arginine-glutamine motif that is thought to function by interacting with G protein catalytic domains to facilitate GTP hydrolysis. These data suggest that the N-terminal tyrosine hydroxyls in Kir3 normally mask the GAP activity and that modification by phosphorylation or phenylalanine substitution reveals the GAP domain. Thus, BDNF activation of trkB could inhibit Kir3 by facilitating channel deactivation.
authors.affiliation: Department of Pharmacology, University of Washington School of Medicine, Seattle, Washington 98195-7280, USA.
authors.forename: Danielle L
authors.initials: DL
authors.lastname: Ippolito
authors.forename: Paul A
authors.initials: PA
authors.lastname: Temkin
authors.forename: Sherri L
authors.initials: SL
authors.lastname: Rogalski
authors.forename: Charles
authors.initials: C
authors.lastname: Chavkin
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D001120:Arginine / Q000737:chemistry; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology; D020134:Catalytic Domain; D004789:Enzyme Activation; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D051960:GTP-Binding Protein alpha Subunit, Gi2; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism / Q000502:physiology*; D005973:Glutamine / Q000737:chemistry; D006868:Hydrolysis; D007700:Kinetics; D008956:Models, Chemical; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000737:chemistry* / Q000378:metabolism; D024661:Potassium Channels, Inwardly Rectifying; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism / Q000502:physiology*; D020710:RGS Proteins / Q000378:metabolism; D018075:RNA, Complementary / Q000378:metabolism; D020813:Receptor, trkB / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry* / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology; D014981:Xenopus
chemical_list: D019208:Brain-Derived Neurotrophic Factor; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011518:Proto-Oncogene Proteins; D020710:RGS Proteins; D018075:RNA, Complementary; D017450:Receptors, Opioid, mu; D005973:Glutamine; C098799:RGS4 protein; D014638:Vanadates; D014443:Tyrosine; D001120:Arginine; D020813:Receptor, trkB; D017027:Protein Tyrosine Phosphatases; D020558:GTP Phosphohydrolases; D051960:GTP-Binding Protein alpha Subunit, Gi2; D019206:GTP-Binding Protein alpha Subunits, Gi-Go