X-RAY INDUCTION OF METHOTREXATE RESISTANCE DUE TO DHFR GENE AMPLIFICATION

被引:67
作者
HAHN, P [1 ]
NEVALDINE, B [1 ]
MORGAN, WF [1 ]
机构
[1] UNIV CALIF SAN FRANCISCO,RADIOBIOL & ENVIRONM HLTH LAB,SAN FRANCISCO,CA 94143
关键词
D O I
10.1007/BF01233191
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The effect of ionizing radiation on methotrexate (MTX) resistance and gene amplification in cultured mammalian cells was investigated. X-irradiation of mouse EMT-6 cells induced cell killing and MTX resistance due to amplification of the dihydrofolate reductase (dhfr) gene in a dose-dependent manner. The highest yields of mutant cells were obtained at approximately D37 (the dose at which 37% of the cells survive), where the frequency of MTX-resistant cells was four- to eightfold over that of the unirradiated population. The proportion of MTX-resistant cells among the survivors increased logarithmically with dose, up to a 1000-fold increase over unirradiated cells at 1000 cGy, the highest dose tested. The induced frequency of MTX resistance after X-irradiation was greater than the induced frequency of 8-azaguanine resistance, which indicates deletion of the hypoxanthine phosphoribosyltransferase gene. Inhibition of poly(ADP-ribose) polymerase by the addition of 3-aminobenzamide before irradiation increased both cell killing and MTX resistance. Metaphase spreads of chromosomes from EMT-6 cells that had been irradiated and subjected to stepwise increases in MTX concentration showed numerous double minutes. Pulsed-field gel electrophoresis of the DNA from cells containing radiation-induced double minutes showed that many copies of the dhfr gene were present on circular DNA molecules of 106, 2× 106, and 3×106 base pairs. These results suggest a relationship between the induction of chromosome aberrations and the induction of gene amplification. © 1990 Plenum Publishing Corporation.
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页码:413 / 423
页数:11
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