Selective overproduction of human dihydrofolate reductase in a methotrexate-resistant human-mouse somatic cell hybrid

The development of methotrexate (MTX) resistance in cultured cells results in increased levels of the drug's target enzyme dihydrofolate reductase (DHFR). Stepwise-selected MTX-resistant sublines originating from an MTX-sensitive human-mouse hybrid expressed elevated DHFR levels and human-DHFR specific gene sequence amplification. By high resolution two-dimensional polyacrylamide gradient electrophoresis, human DHFR was shown to be selectively overproduced in VB2a-100 MTX-resistant cells whereas mouse DHFR protein "spots" present in MTX-sensitive parental hybrid were absent in these cells exhibiting 100 microM MTX resistance. These findings and those in a parallel study indicate that concurrent with overproduction of human DHFR and amplification DHFR sequences in VB2a-100, a loss of mouse-specific DHFR gene sequences occurred.