Eastman H B, Swick A G, Schmitt M C, Azizkhan J C
Lineberger Cancer Research Center, University of North Carolina, Chapel Hill 27599-7295.
Proc Natl Acad Sci U S A. 1991 Oct 1;88(19):8572-6. doi: 10.1073/pnas.88.19.8572.
Dihydrofolate reductase (DHFR; EC 1.5.1.3) is required in folate metabolism for the synthesis of purines, thymidine, and glycine. Although there have been several reports of induction of DHFR enzyme by methotrexate (MTX), a drug that competitively inhibits DHFR, there are no studies reported that examine the effect of MTX on DHFR gene transcription. We have examined the effect of MTX and other inhibitors of DNA synthesis on DHFR transcription using a transient expression assay. MTX stimulates transient expression in a concentration-dependent manner from a hamster DHFR promoter construct containing 150 base pairs 5' to the start of transcription. Addition of either tetrahydrofolate or hypoxanthine plus thymidine prevents the promoter induction in response to MTX, suggesting that stimulation by MTX results from inhibition of these metabolites. Furthermore, two other antimetabolic drugs--fluorodeoxyuridine and hydroxyurea--also stimulate the DHFR promoter in a concentration-dependent manner. In contrast, aphidicolin, which blocks cell growth through inhibition of DNA polymerase alpha, has no effect on the DHFR promoter. The potential relevance of these results to cross-resistance to chemotherapeutic agents and to the process of gene amplification is discussed.
二氢叶酸还原酶（DHFR；EC 1.5.1.3）在叶酸代谢中对于嘌呤、胸苷和甘氨酸的合成是必需的。尽管已有数篇关于甲氨蝶呤（MTX）诱导DHFR酶的报道，甲氨蝶呤是一种竞争性抑制DHFR的药物，但尚无研究报道检测MTX对DHFR基因转录的影响。我们使用瞬时表达分析检测了MTX和其他DNA合成抑制剂对DHFR转录的影响。MTX以浓度依赖的方式刺激来自包含转录起始点上游150个碱基对的仓鼠DHFR启动子构建体的瞬时表达。添加四氢叶酸或次黄嘌呤加胸苷可阻止响应MTX的启动子诱导，这表明MTX的刺激是由于这些代谢物的抑制所致。此外，另外两种抗代谢药物——氟脱氧尿苷和羟基脲——也以浓度依赖的方式刺激DHFR启动子。相比之下，通过抑制DNA聚合酶α来阻断细胞生长的阿非科林对DHFR启动子没有影响。讨论了这些结果与化疗药物交叉耐药性以及基因扩增过程的潜在相关性。
