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Anti folate resistant chinese hamster cells messenger rna directed over production of multiple di hydro folate reductases from a series of independently derived sublines containing amplified di hydro folate reductase genes



Anti folate resistant chinese hamster cells messenger rna directed over production of multiple di hydro folate reductases from a series of independently derived sublines containing amplified di hydro folate reductase genes



Journal of Biological Chemistry 257(21): 12939-12949



Purification of the dihydrofolate reductases overproduced by 16 independently-derived antifolate-resistant sublines of Chinese hamster lung cells and their drug-sensitive parental cell line, DC-3F, has confirmed the original observation that 2 MW classes of this enzyme, i.e, MW = 21,000 and MW = 20,000, can be overproduced by drug-resistant cells. Of the 16 drug-resistant sublines analyzed, 12 overproduce the 21,000-dalton enzyme, while 4 overproduce the 20,000-dalton enzyme. Both MW classes are found in the drug-sensitive parental line. Two-dimensional polyacrylamide gel electrophoresis of the 2 MW classes of enzyme showed that each is composed of 2 isoelectric forms, and that the relative distributins of the isoelectric forms within a MW class are maintained during overproduction. In vitro translation experiments demonstrated that the synthesis of these isoelectric forms and their relative distributions are mediated by mRNA. Evidence is presented for the existence of 2 sets of 3 dihydrofolate reductase mRNA in overproducing sublines. Both sets, within a subline, encode the same MW class of dihydrofolate reductase, while each set directs the synthesis of only one of the isoelectric forms within that class. Although the genetic basis for these observations is not yet understood, it is clear that the production of dihydrofolate reductase by Chinese hamster lung cells is complex and probably involves the expression of multiple dihydrofolate reductase genes and/or alleles.

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Accession: 004757389

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