Cycloheximide efflux in antibiotic-adapted cells of the fungus Mucor racemosus

Abstract

Mucor racemosus cells adapted to either cycloheximide or trichodermin were approximately 40-fold more resistant to cycloheximide than nonadapted cells. Ribosomes isolated from adapted and nonadapted cells were equally sensitive to cycloheximide in an in vitro poly(U) translation assay. There was no detectable modification of cycloheximide by adapted cells. Uptake of drug by nonadapted and adapted cells was characterized by a rapid initial accumulation during the first 2 min of incubation with [3H]cycloheximide, followed by a steady-state intracellular drug concentration well below that of the medium. The steady-state drug concentration was approximately 10-fold lower in adapted cells than in nonadapted cells. Treatment of cells with sodium azide or dinitrophenol abolished the difference between uptake of drug by nonadapted and adapted cells and resulted in intracellular drug levels equal to that of the medium. Direct efflux measurements showed that adapted cells loaded with cycloheximide were able to excrete the drug far more rapidly than nonadapted cells. These results suggest that both nonadapted and adapted cells possess an energy-dependent efflux mechanism for transporting cycloheximide and that resistance in adapted cells is due to increased efficiency of transport.