Characterization of Cytochrome P450 and a Putative Cytochrome P450 Gene in Drosophila melanogaster

Abstract

Cytochrome P450 was examined in both insecticide resistant and insecticide susceptible strains of Drosophila melanogaster. Much higher levels were observed in the resistant strain IIID when compared to the susceptible strain Canton S. This increase appeared to be the result of an overproduction of a few existing forms. Two heme-staining microsomal proteins found in strain IIID were identified as putative cytochrome P450 isozymes. Polyclonal antibodies produced against these two proteins were used in the immunoanalysis of microsomal proteins from both strains. A lambda gtll cDNA expression vector library was created by inserting cDNA fragments from a Drosophila lambda gt10 library into lambda gtll arms. The library was screened with the polyclonal antiserum. Three clones were isolated, of which one, gtll-Al, was most highly reactive with the antiserum. Analysis of the gtll-Al lysogen indicated a 130 kd fusion protein was produced of which 16 kd was coded for by the cDNA insert. A .5 kb cDNA insert was isolated from the clone as part of a 1.5 kb Kpnl/EcoRl fragment and was used in the analysis of Drosophila genomic DNA and total RNA. Southern analysis revealed an EcoRl polymorphism existed between strain IIID and Canton S. RNA analysis suggested strain IIID produced more coding message for the Al insert in the larval and adult stages than did Canton S.