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.
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Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada.
Andrulis
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D000599:Amino Acids, Diamino / Q000494:pharmacology*; D000818:Animals; D001217:Aspartate-Ammonia Ligase / Q000096:biosynthesis* / Q000235:genetics; D015139:Blotting, Southern; D015153:Blotting, Western; D004171:Diploidy; D004351:Drug Resistance / Q000235:genetics; D005784:Gene Amplification; D051379:Mice; D012333:RNA, Messenger / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D000599:Amino Acids, Diamino; D012333:RNA, Messenger; D001217:Aspartate-Ammonia Ligase; C006440:albizziin