Turner P R, Jaffe L A, Fein A
J Cell Biol. 1986 Jan;102(1):70-6. doi: 10.1083/jcb.102.1.70.
To investigate the roles of inositol 1,4,5-trisphosphate (InsP3) and guanyl nucleotide binding proteins (G-proteins) in the transduction mechanism coupling fertilization and exocytosis of cortical vesicles in sea urchin eggs, we microinjected InsP3 and guanyl nucleotide analogs into eggs of Lytechinus variegatus. Injection of 28 nM InsP3 caused exocytosis. However, if the egg was first injected with EGTA ([Cai] less than or equal to 0.1 microM; EGTA = 1.6 mM), InsP3 injection did not cause exocytosis, supporting the hypothesis that InsP3 acts by causing a rise in intracellular free calcium. Injection of 28 microM guanosine-5'-0-(3-thiotriphosphate) (GTP-gamma-S), a hydrolysis-resistant analog of GTP, caused exocytosis, but exocytosis did not occur if the egg was pre-injected with EGTA. Injection of 3 mM guanosine-5'-0-(2-thiodiphosphate) (GDP-beta-S), a metabolically stable analog of GDP, prevented sperm from stimulating exocytosis. However, injection of GDP-beta-S did not prevent the stimulation of exocytosis by InsP3. These results suggested the following sequence of events. The sperm activates a G-protein, which stimulates production of InsP3. InsP3 elevates intracellular free calcium, which causes exocytosis.
为了研究肌醇1,4,5 - 三磷酸（InsP3）和鸟苷酸结合蛋白（G蛋白）在海胆卵受精与皮质囊泡胞吐作用偶联的转导机制中的作用，我们将InsP3和鸟苷酸类似物显微注射到多色灯塔海胆的卵中。注射28 nM的InsP3会引起胞吐作用。然而，如果先向卵中注射乙二醇双四乙酸（[Ca²⁺]≤0.1 μM；乙二醇双四乙酸 = 1.6 mM），再注射InsP3则不会引起胞吐作用，这支持了InsP3通过引起细胞内游离钙升高而起作用的假说。注射28 μM鸟苷 - 5'-O -（3 - 硫代三磷酸）（GTP - γ - S），一种GTP的抗水解类似物，会引起胞吐作用，但如果卵预先注射了乙二醇双四乙酸则不会发生胞吐作用。注射3 mM鸟苷 - 5'-O -（2 - 硫代二磷酸）（GDP - β - S），一种代谢稳定的GDP类似物，可阻止精子刺激胞吐作用。然而，注射GDP - β - S并不能阻止InsP3对胞吐作用的刺激。这些结果提示了以下事件顺序。精子激活一种G蛋白，该G蛋白刺激InsP3的产生。InsP3升高细胞内游离钙，进而引起胞吐作用。
