Overexpression of asparagine synthetase in albizziin-resistant murine diploid embryonic stem cells.

Gene amplification is commonly observed in primary tumors and established drug-resistant cell lines, both of which are generally aneuploid. However, this process is undetectable (frequency < 10(-9) in normal diploid mammalian cell lines. To investigate whether gene amplification can occur in pluripotent diploid cells, we have selected drug-resistant mutants of mouse embryonic stem (ES) cells. We had previously found that Chinese hamster ovary (CHO) and human cell lines selected in albizziin (Alb), an amino acid analog of L-glutamine, overexpress asparagine synthetase (AS) due to gene amplification. The same drug selection system was applied to ES cells to isolate single-step and multistep drug-resistant mutants. Albizziin-resistant ES cells exhibited elevated levels of AS; however, drug resistance in ES cells was associated with mRNA overexpression without gene amplification. AS gene amplification was observed in only one drug-resistant cell line and was preceded by AS mRNA overexpression. Gene amplification in the latter coincided with the loss of the pluripotent nature of the ES cells.