Expression of carbamoyl phosphate synthetase I and ornithine transcarbamoylase genes in Chinese hamster ovary dhfr-cells decreases accumulation of ammonium ion in culture media

Ammonium ion accumulation in mammalian cell culture media causes toxicity w hich inhibits cell growth and productivity, To reduce the level of the accu mulated ammonium ion, carbamoyl phosphate synthetase I (CPS I) and ornithin e transcarbamoylase (OTC) were used, which catalyze the first and second st eps of the urea cycle in the liver. To examine the effects of overexpressed CPS I and OTC genes on the concentration of the ammonium ion in culture me dia, the two genes were introduced into Chinese hamster ovary (CHO) dhfr-ce lls. The CPS I expressing cell lines (CPS I-CHO) and both CPS I and OTC exp ressing cell lines (CPS I/OTC-CHO) were confirmed at the mRNA level and ana lyzed in terms of the cell growth and the accumulation of ammonium ion in c ulture media. The accumulation of ammonium ion was approximate to 25-33% le ss in CPS I/OTC-CHO than in either CPS I-CRO or the vector-control cell lin es. Interestingly however, the cell growth was approximate to 15-30% faster in both CPS I-CHO and CPS I/OTC-CHO than in the control cell lines. Forced expression of urea cycle enzymes in the CHO cells revealed that both the e xpression of CPS I and OTC can reduce the accumulation of ammonium ion in t he culture media. (C) 2000 Elsevier Science B.V. All rights reserved.