Elevated amounts of methotrexate-binding protein,different from normal dihydrofolate reductase,in a petunia MTXR-cell line |
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Authors: | Rivka Barg Meir Perl Jacques S Beckmann |
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Institution: | (1) Department of Plant Genetics and Breeding, Agricultural Research Organization, the Volcani Center, 50250 Bet Dagan, Israel;(2) Department of Medicinal and Spice Crops, Agricultural Research Organization, the Volcani Center, 50250 Bet Dagan, Israel |
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Abstract: | Summary A petunia cell line, 1ECB, was previously isolated by the stepwise selection procedure, for resistance to methotrexate (MTX), an antimetabolite for the enzyme dihydrofolate reductase (DHFR). Using ammonium sulfate precipitates of cell lysates of cell line 1ECB and its parental cell line (WT), it was found that the mutant has an increase of 400 fold in 3H-MTX binding capacity and a decrease in the affinity for MTX binding, at two orders of magnitude, in comparison with the WT. In addition, the DHFR specific activity in the mutant increased only moderately (5- to 10-fold), this activity is extremely sensitive to MTX inhibition as compared to the WT. It is evident that the MTX resistance of line 1ECB results mainly from overproduction of an MTX-binding protein which differs from the WT DHFR by four biochemical criteria. This protein may serve as a trap for the excess amounts of MTX to which the cells are exposed. |
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Keywords: | amplification DHFR MTX-binding protein MTX-resistance petunia cell suspension stepwise selection |
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