OVERPRODUCTION OF A BIFUNCTIONAL THYMIDYLATE SYNTHETASE-DIHYDROFOLATE REDUCTASE AND DNA AMPLIFICATION IN METHOTREXATE-RESISTANT LEISHMANIA-TROPICA
OVERPRODUCTION OF A BIFUNCTIONAL THYMIDYLATE SYNTHETASE-DIHYDROFOLATE REDUCTASE AND DNA AMPLIFICATION IN METHOTREXATE-RESISTANT LEISHMANIA-TROPICA
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DOI：
10.1073/pnas.80.8.2132
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发表时间：
1983-01-01
期刊：
影响因子：
--
通讯作者：
SANTI, DV
中科院分区：
文献类型：
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作者：
CODERRE, JA;BEVERLEY, SM;SANTI, DV
L.开发了对氨甲蝶呤（一种二氢叶酸还原酶抑制剂）具有高度抗性的热带前鞭毛体。对1 mM甲氨蝶呤具有抗性的生物体在与胸苷酸合成酶相关的二氢叶酸还原酶中具有40倍的增加，并且它们含有可以在限制性内切酶的染色凝胶上直接可视化的DNA扩增区域。这些生物体中扩增的DNA长度约为56个内切酶，拷贝数约为野生型生物体的80倍，约占核DNA的10%。当抗甲氨蝶呤L.当在无药物培养基中繁殖热带假单胞菌时，二氢叶酸还原酶-胸苷酸合成酶蛋白和扩增的DNA以平行的方式减少，直到它们与野生型生物体中的水平不可区分。当这些明显的回复突变体再次用1 mM甲氨蝶呤挑战时，酶的过度生产和DNA扩增迅速发生。在哺乳动物细胞中，寄生原生动物的抗药性似乎是由扩增介导的。
L. tropica promastigotes that are highly resistant to methotrexate, a dihydrofolate reductase inhibitor, were developed. Organisms resistant to 1 mM methotrexate have a 40-fold increase in dihydrofolate reductase which is associated with thymidylate synthetase, and they contain amplified regions of DNA that may be directly visualized on stained gels of restriction digests. The amplified DNA in these organisms is about 56 kilobases in length, has a copy number about 80-fold higher than that of wild-type organisms, and constitutes about 10% of the nuclear DNA. When the methotrexate-resistant L. tropica are propagated in drug-free medium, the dihydrofolate reductase-thymidylate synthetase protein and the amplified DNA decrease in a parallel fashion until they are indistinguishable from the levels in wild-type organisms. When these apparent revertants are again challenged with 1 mM methotrexate, enzyme overproduction and DNA amplification occur rapidly. As in mammalian cells, it appears that drug resistance in parasitic protozoa may be mediated by amplification.