Phosphatidylinositol 3,4,5-trisphosphate and Ca2+/calmodulin competitively bind to the regulators of G-protein-signalling (RGS) domain of RGS4 and reciprocally regulate its action.
RGS (regulators of G-protein signalling) are a diverse group of proteins, which accelerate intrinsic GTP hydrolysis on heterotrimeric G-protein a subunits. They are involved in the control of a physiological behaviour known as 'relaxation' of G-protein-gated K+ channels in cardiac myocytes. The GTPase-accelerating activity of cardiac RGS proteins, such as RGS4, is inhibited by PtdIns(3,4,5)P3 (phosphatidylinositol 3,4,5-trisphosphate) and this inhibition is cancelled by Ca2+/calmodulin (CaM) formed during membrane depolarization. G-protein-gated K+ channel activity decreases on depolarization owing to the facilitation of GTPase-activating protein activity by RGS proteins and vice versa on hyperpolarization. The molecular mechanism responsible for this reciprocal control of RGS action by PtdIns(3,4,5)P3 and Ca2+/CaM, however, has not been fully elucidated. Using lipid-protein co-sedimentation assay and surface plasmon resonance measurements, we show in the present study that the control of the GTPase-accelerating activity of the RGS4 protein is achieved through the competitive binding of PtdIns(3,4,5)P3 and Ca2+/CaM within its RGS domain. Competitive binding occurs exclusively within the RGS domain and involves a cluster of positively charged residues located on the surface opposite to the Ga interaction site. In the RGS proteins conserving these residues, the reciprocal regulation by PtdIns(3,4,5)P3 and Ca2+/CaM may be important for their physiological regulation of G-protein signalling.
Department of Pharmacology, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871, Japan.
Masaru
M
Ishii
Satoru
S
Fujita
Mitsuhiko
M
Yamada
Yukio
Y
Hosaka
Yoshihisa
Y
Kurachi
D000109:Acetylcholine / Q000494:pharmacology; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002147:Calmodulin / Q000378:metabolism* / Q000494:pharmacology; D002417:Cattle; D002942:Circular Dichroism; D004553:Electric Conductivity; D004578:Electron Spin Resonance Spectroscopy; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D015221:Potassium Channels / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D020710:RGS Proteins / Q000737:chemistry* / Q000378:metabolism*; D013379:Substrate Specificity; D014981:Xenopus
D002147:Calmodulin; D044385:GTP-Binding Protein alpha Subunits; D018129:Phosphatidylinositol Phosphates; D015221:Potassium Channels; D020710:RGS Proteins; C060974:phosphatidylinositol 3,4,5-triphosphate; C098799:RGS4 protein; D008239:Lysine; D000109:Acetylcholine; D002118:Calcium