PHOSPHATIDYL-INOSITOL PHOSPHOLIPASE-C IN SQUID PHOTORECEPTOR MEMBRANEIS ACTIVATED BY STABLE METARHODOPSIN VIA GTP-BINDING PROTEIN, GQ

Phosphatidyl inositol-phospholipase C (PI-PLC) in squid retina was stu died by immunoblotting and its activities were determined using [H-3]p hosphatidyl inositol bisphosphate ([H-3]PIP2) as substrate. PI-PLC act ivity was found mostly in soluble fraction when the retina homogenate was treated with 400 mM KCl, but was associated with rhabdomal membran es under low salt conditions (20 mM Hepes). A protein with apparent mo lecular mass of 130 kD was recognized by an antibody against PLC beta 4/norp A in both 400 mM KCl soluble and rhabdomal membrane fractions. A 42 kD protein recognized by antibody against the C-terminus of Gq al pha was also present in these two fractions. GTP gamma S stimulated on ly the PI-PLC activity associated with membrane and was magnesium depe ndent. PI-PLC activity was found to be (i) highly dependent upon calci um concentrations, (ii) enhanced by GTP but not by other nucleotides, and (iii) significantly stimulated by light at lower concentrations of GTP gamma S. The stimulation by light was still observed when irradia ted membrane was incubated at 10 degrees C for 10 min and then mixed w ith GTP gamma S. These results suggest that stable metarhodopsin stimu lates a PLC beta 4/norp A-like enzyme via a G-protein, Gq.