Mechanism of action of forskolin on adenylate cyclase: Effect on bovine sperm complemented with erythrocyte membranes

The mechanism of action of forskolin stimulation of adenylate cyclase was investigated by examining its effects on the enzyme's Mg²⁺ activated catalytic unit (C) from bovine sperm, both preceding and following complementation with human erythrocyte membranes as a source of guanine nucleotide regulatory protein (N). Prior to complementation, sperm C was not activated by either NaF (10 mM) or 5′-guanylyl-β-γ-imidodiphosphate (Gpp(NH)p, 10 μM), suggesting that functional N was not present in this preparation. Forskolin (100 μM) was also without effect on C. Following complementation of the sperm membranes with those of erythrocytes, Mg²⁺-dependent sensitivity to forskolin, NaF, and Gpp(NH)p was imparted to C. Our findings are incompatible with the current hypothesis that forskolin stimulates adenylate cyclase by direct activation of C. Rather, the data suggest that the activation process occurs through an effect on N or by augmentation of the interaction between the components of the adenylate cyclase complex.