Inhibition of apoptosis by overexpressing Bcl-2 enhances gene amplification by a mechanism independent of aphidicolin pretreatment.
Inhibition of apoptosis by overexpressing Bcl-2 enhances gene amplification by a mechanism independent of aphidicolin pretreatment.
复制标题
通过过表达 Bcl-2 抑制细胞凋亡，通过独立于阿非迪霉素预处理的机制增强基因扩增。
DOI：
10.1073/pnas.93.8.3394
复制
发表时间：
1996
影响因子：
11.1
通讯作者：
Schimke,RT
中科院分区：
文献类型：
--
作者：
Yin,DX;Schimke,RT
为了研究细胞凋亡对基因扩增的影响，我们在生长培养基中构建了四环素可控制bcl-2 （BCL2）表达的HeLa S3细胞株。当使用三甲氨蝶呤选择细胞时，Bcl-2表达的诱导导致细胞凋亡的暂时延迟，并导致抗性菌落的频率增加约3倍。这种抗性是由于二氢叶酸还原酶基因的扩增。无论诱导还是抑制Bcl-2，从聚集的抗性菌落中生长出来的细胞对三甲氨蝶呤的抗性水平都是相同的，这与Bcl-2通过促进基因扩增而不是抗性机制本身起作用的理论是一致的。用阿霉素预处理细胞是另一种增加基因扩增频率的方法。当表达Bcl-2的细胞经阿菲迪克林预处理后，基因扩增频率的增加大致是阿菲迪克林预处理或Bcl-2单独表达增加的产物，说明Bcl-2增加基因扩增的机制独立于阿菲迪克林预处理。这些结果与基因扩增在药物诱导的细胞周期扰动中以更高频率发生的概念是一致的。Bcl-2在扰动过程中通过延长细胞的存活时间，明显增加了扩增集落的选择数量。
To study the effect of apoptosis on gene amplification, we have constructed HeLa S3 cell lines in which the expression of bcl-2 (BCL2) can be controlled by tetracycline in the growth medium. Induction of Bcl-2 expression caused a temporary delay of apoptosis and resulted in roughly a 3-fold increase in the frequency of resistant colonies when cells were selected with trimetrexate. This resistance was due to amplification of the dihydrofolate reductase gene. Cells grown out of the pooled resistant colonies retained the same level of resistance to trimetrexate whether Bcl-2 was induced or repressed, consistent with the theory that Bcl-2 functions by facilitating gene amplification, rather than being the resistance mechanism per se. Pretreating cells with aphidicolin is another method to increase gene amplification frequency. When Bcl-2-expressing cells were pretreated with aphidicolin, the resulting increase in gene amplification frequency was approximately the product of the increases caused by aphidicolin pretreatment or Bcl-2 expression alone, indicating that Bcl-2 increases gene amplification through a mechanism independent of that of aphidicolin pretreatment. These results are consistent with the concept that gene amplification occurs at a higher frequency during drug-induced cell cycle perturbation. Bcl-2 evidently increases the number of selected amplified colonies by prolonging cell survival during the perturbation.