Selection of Chinese hamster ovary cells (CHO-K1) with reduced glutamate and aspartate uptake.

ABSTRACT

Transport of L-[3H]glutamate into Chinese hamster ovary cells (CHO-K1) was characterized and the results used to design a tritium suicide selection for cells with transport defects. Replicas of surviving colonies on polyester cloth disks were screened by autofluorography for reduced uptake and two mutant clones, Ed-A1 and Ed-B8, were obtained. Uptake of glutamate through a sodium-dependent system in both mutants was characterized by significant reductions in Vmax and increases in Km compared to parental cells, but their response to removal of extracellular sodium differed, suggesting distinct mutations in the two lines. The Vmax of aspartate uptake through this system was reduced in both mutants, to one-ninth in the case of Ed-B8. Glutamate uptake through a sodium-independent system was not altered in either mutant. Surprisingly, acid-soluble intracellular pools of several amino acids were higher in both mutants.