Handling of L-[35S]cystine by cysteamine-pretreated cystinotic and normal fibroblasts.

In short incubations with 0.1 mM L-[35S]cystine in phosphate-buffered saline medium, and long incubations with label in complete minimum Eagle's medium with Earle salts, cystine-depleted cystinotic cells reaccumulate labeled cystine more rapidly than pretreated normal cells. Cysteamine pretreatment of both normal and cystinotic cells resulted in an initial increased conversion of exogenous cystine to intracellular cysteine. In 24-h incubations in complete medium, cysteamine-pretreated cells showed enhanced conversion of 0.1 mM L-[35S]cystine to cysteine and reduced glutathione. Addition of cycloheximide to the incubation media decreased the incorporation of 35S into cellular protein by more than 90% but did not affect the accumulation of intracellular labeled cystine in cystinotic cells. Therefore, the incorporation and release of cystine from protein is not an obligatory source of accumulated cystine and we speculate that there may be early extralysosomal entrapment of cystine in cystinotic cells.