Cysteine control over glutathione homeostasis in Chinese hamster fibroblasts overexpressing a γ‐glutamylcysteine synthetase activity

γ‐Glutamylcysteine synthetase (GCS) catalyses the first step of glutathione (GSH) biosynthesis and is considered to be the rate‐limiting step of this pathway. In several experimental systems, GCS overexpression has been associated with GSH pool expansion and drug resistance. In this report, we describe a mutant line of Chinese hamster fibroblasts that overexpress this activity by 4–5 times, due to the amplification of the gene encoding the catalytic subunit of GCS. These mutant cells contained a wild‐type steady‐state level of GSH and, after depletion, synthesized GSH at the same rate as wild‐type cells because their rate of endogenous production of cysteine was limiting. An exogenous supply of cysteine expanded the pool of GSH in mutant cells by 80% but did not increase that of wild‐type cells, and, in GSH‐depleted cells, increased the rate of GSH biosynthesis by eight and 35‐times in wild‐type and mutant cells, respectively. These experiments indicated that GCS overexpression had no consequence on the metabolism of GSH, unless a supply of cysteine was provided. Mutant cells were not resistant to cisplatin or nitrogen mustard.