Transport of L-cystine by cultivated skin fibroblasts of normal subjects and patients with cystinosis

Uptake of L-cystine at the plasma membrane of fibroblasts derived from normal and cystinotic subjects was studied. L-Cystine accumulation after a 20-min period was increased in cystinotic fibroblasts incubated in 0.08 mM L-cystine. This effect appeared to be concentration-dependent since accumulation after 20 min at 0.004 mM concentration was decreased in cystinotic cells. Kinetic data suggested that at least two nondiffusional saturable processes with widely different substrate affinities mediate initial L-cystine uptake in skin fibroblasts. In addition, the transport process with high affinity for L-cystine may itself be a two-component system, as suggested by (/) additive inhibtory effect of other neutral amino acids, and (2) preincubation studies in which preincubation with cystathionine enhanced subsequent L-cystine uptake, whereas preincubation with other neutral amino acids depressed subsequent uptake. Affinity constants and maximal velocities of initial uptake did not appear to be altered in cells derived from patients with cystinosis. After 60-sec incubation with L-[35S]cystine, cystinotic cells retained more label as cystine than did normal cells at each concentration studied. These data indicate that initial L-cystine uptake in fibroblasts of patients with cystinosis proceeds at a normal rate by means of all transport systems currently shown to be present in normal cells. L-Cystine may accumulate in cells of patients with cystinosis because of an alteration in a transport system for L-cystine which is, as yet, undescribed. Alternatively, L-cystine accumulation may occur because of a transport defect at the lysosomal membrane which is not expressed at the plasma membrane.