Effects of ionophores on vitellogenin uptake by Hyalophora oocytes

Oocytes of Hyalophora cecropia that were incubated in vitro with ³⁵S]vitellogenin incorporated label within 10 min into an intermediate-density compartment identified by sucrose density gradient centrifugation. During a subsequent 20-min chase this presumptive endosomal label was transferred to a compartment with the higher density of protein yolk spheres. When vitellogenin uptake was inhibited by 10 μM nigericin or monensin, or 50 μM carbonyl cyanide m-cholorophenylhydrazone, a somewhat larger and more focused peak of label accumulated in the endosome region of the gradient, and the transfer of this label to the yolk spheres was blocked. Valinomycin, at concentrations as high as 100 μM, did not inhibit uptake or processing, even though successful insertion into the oocyte membrane could be demonstrated by the effects of this ionophore on the membrane potential and K⁺ permeability of the follicle. Inhibition of processing by nigericin and monensin is consistent with a model of endocytosis in which the ionophores prevent acidification of the endosomes by promoting H⁺-K⁺ exchange with the cytoplasm. Several alternative possibilities were ruled out by physiological analyses entailing the measurement of cytoplasmic pH and membrane potentials.