Endocytosis in Dictyostelium discoideum amoebae: Inhibition by cyclohexlmide and other inhibitors of protein synthesis

Abstract

ABSTRACT Cycloheximide, an inhibitor of protein synthesis, inhibited endocytosis pathways in the vegetative amoebae of the cellular slime mold Dictyostelium discoideum. Half-maximal inhibition of fluid-phase pinocytosis in nutrient medium or in buffer occurred at 0.08 mM or 0.35 mM cycloheximide, respectively. Other inhibitors of eucaryotic protein synthesis inhibited fluid-phase pinocytosis, with the following IC50 in nutrient medium: 0.15 mM anisomycin, ImM emetine, 0.25 mM puromycin. Kinetic studies showed that the inhibitory effects of cycloheximide occurred rapidly, with a half-time of less than 10 min, and were fully reversible upon removal of the drug. Both fluid-phase pinocytic influx and efflux were inhibited by cycloheximide, which furthermore prevented the vanadate-induced efflux of fluorescein-labeled dextran, the fluid-phase marker used in these experiments. The massive secretion of lysosomal enzymes: acid phosphatase, N-acetylhexosaminidase or α-mannosidase, which follows overloading of the endosomal compartment by sucrose, was also inhibited by cycloheximide. All these phenomena were unimpaired by cycloheximide in the cycloheximide-resistant (cycAl) mutant strain X6.