G alpha(q/11) coupled to mammalian phospholipase C beta 3-like enzyme mediates the ginsenoside effect on Ca(2+)-activated Cl(-) current in the Xenopus oocyte.
Recently we demonstrated that ginsenosides, the active ingredients of Panax ginseng, enhanced Ca(2+)-activated Cl(-) current in the Xenopus oocyte through a signal transduction mechanism involving the activation of pertussis toxin-insensitive G protein and phospholipase C (PLC). However, it has not yet been determined precisely which G protein subunit(s) and which PLC isoform(s) participate in the ginsenoside signaling. To provide answers to these questions, we investigated the changes in ginsenoside effect on the Cl(-) current after intraoocyte injections of the cRNAs coding various G protein subunits, a regulator of G protein signaling (RGS2), and G beta gamma-binding proteins. In addition, we examined which of mammalian PLC beta 1-3 antibodies injected into the oocyte inhibited the action of ginsenosides on the Cl(-) current. Injection of G alpha(q) or G alpha(11) cRNA increased the basal Cl(-) current recorded 48 h after, and it further prevented ginsenosides from enhancing the Cl(-) current, whereas G alpha(i2) and G alpha(oA) cRNA injection had no significant effect. The changes following G alpha(q) cRNA injection were prevented when G beta(1)gamma(2) and G alpha(q) subunits were co-expressed by simultaneous injection of the cRNAs coding these subunits. Injection of cRNA coding G alpha(q)Q209L, a constitutively active mutant that does not bind to G beta gamma, produced effects similar to those of G alpha(q) cRNA injection. The effects of G alpha(q)Q209L cRNA injection, however, were not prevented by co-injection of G beta(1)gamma(2) cRNA. Injection of the cRNA coding RGS2, which interacts most selectively with G alpha(q/11) among various identified RGS isoforms and stimulates the hydrolysis of GTP to GDP in active GTP-bound G alpha subunit, resulted in a severe attenuation of ginsenoside effect on the Cl(-) current. Finally, antibodies against PLC beta 3, but not -beta 1 and -beta 2, markedly attenuated the ginsenoside effect examined at 3-h postinjection. These results suggest that G alpha(q/11) coupled to mammalian PLC beta 3-like enzyme mediates ginsenoside effect on Ca(2+)-activated Cl(-) current in the Xenopus oocyte.
Choi
S
S
National Research Laboratory for the Study of Ginseng Signal Transduction and Department of Physiology, College of Veterinary Medicine, Chonnam National University, Kwangju 500-757 Korea.
Kim
H J
HJ
Ko
Y S
YS
Jeong
S W
SW
Kim
Y I
YI
Simonds
W F
WF
Oh
J W
JW
Nah
S Y
SY
D000818:Animals; D020013:Calcium Signaling / Q000502:physiology*; D002491:Central Nervous System Agents / Q000494:pharmacology; D018118:Chloride Channels / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D019204:GTP-Binding Proteins / Q000378:metabolism; D036145:Ginsenosides; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D008845:Microinjections; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D005894:Panax / Q000737:chemistry; D018408:Patch-Clamp Techniques; D054799:Phospholipase C beta; D020033:Protein Isoforms; D012313:RNA / Q000378:metabolism; D012503:Saponins / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000378:metabolism*; D014981:Xenopus / Q000502:physiology
D002491:Central Nervous System Agents; D018118:Chloride Channels; D036145:Ginsenosides; D007527:Isoenzymes; D020033:Protein Isoforms; D012503:Saponins; D012313:RNA; D010738:Type C Phospholipases; D054799:Phospholipase C beta; D019204:GTP-Binding Proteins; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins