Fig. 4. Mating input-mediated conformational activation of Ste5.

(A) Simple AND-gate model for specific mating pathway activation. Non-mating inputs that activate the shared MAPKKK do not activate Fus3.

(B) Revised molecular model for mating pathway activation mediated by the Ste5 scaffold protein. Mating pheromone (α-factor) activates a heterotrimeric G protein, leading to release of the Gβγ subunit from Gα and recruitment of Ste5 to free Gβγ at the membrane (21). Membrane recruitment triggers activation of the MAPKKK Ste11 and PH domain binding to PIP₂, leading to release of the VWA domain and relief of autoinhibition.

(C) Fus3 activation in vivo when kinase cascade activation is decoupled from the mating signal (α-factor), measured by protein immunoblotting. See fig. S15 for additional Ste11 alleles and controls.