Wei C, Andrulis I L
Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada.
Somat Cell Mol Genet. 1993 Jul;19(4):321-30. doi: 10.1007/BF01232745.
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.
基因扩增在原发性肿瘤和已建立的耐药细胞系中普遍存在，这两者通常都是非整倍体。然而，在正常二倍体哺乳动物细胞系中，这个过程是无法检测到的（频率<10^(-9)）。为了研究基因扩增是否会在多能二倍体细胞中发生，我们筛选了小鼠胚胎干细胞（ES细胞）的耐药突变体。我们之前发现，在L-谷氨酰胺的氨基酸类似物白雀珊瑚碱（Alb）中筛选得到的中国仓鼠卵巢（CHO）细胞系和人类细胞系，由于基因扩增而过度表达天冬酰胺合成酶（AS）。同样的药物筛选系统应用于ES细胞，以分离单步和多步耐药突变体。对白雀珊瑚碱耐药的ES细胞表现出AS水平升高；然而，ES细胞中的耐药性与mRNA过度表达相关，而没有基因扩增。仅在一个耐药细胞系中观察到AS基因扩增，并且在AS mRNA过度表达之前出现。后者中的基因扩增与ES细胞多能性的丧失同时发生。
