Analysis of a Candida albicans gene that encodes a novel mechanism for resistance to benomyl and methotrexate.
The pathogenic yeast, Candida albicans, is insensitive to the anti-mitotic drug, benomyl, and to the dihydrofolate reductase inhibitor, methotrexate. Genes responsible for the intrinsic drug resistance were sought by transforming Saccharomyces cerevisiae, a yeast sensitive to both drugs, with genomic C. albicans libraries and screening on benomyl or methotrexate. Restriction analysis of plasmids isolated from benomyl- and methotrexate-resistant colonies indicated that both phenotypes were encoded by the same DNA fragment. Sequence analysis showed that the fragments were nearly identical and contained a long open reading frame of 1694 bp (ORF1) and a small ORF of 446 bp (ORF2) within ORF1 on the opposite strand. By site-directed mutagenesis, it was shown that ORF1 encoded both phenotypes. The protein had no sequence similarity to any known proteins, including beta-tubulin, dihydrofolate reductase, and the P-glycoprotein of the multi-drug resistance family. The resistance gene was detected in several C. albicans strains and in C. stellatoidea by DNA hybridization and by the polymerase chain reaction.
Fling
M E
ME
Department of Molecular Genetics and Microbiology, Burroughs Wellcome Co., Research Triangle Park, NC 27709.
Kopf
J
J
Tamarkin
A
A
Gorman
J A
JA
Smith
H A
HA
Koltin
Y
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D001542:Benomyl / Q000494:pharmacology*; D002176:Candida albicans / Q000187:drug effects* / Q000235:genetics; D003062:Codon / Q000235:genetics; D004352:Drug Resistance, Microbial / Q000235:genetics*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics; D005800:Genes, Fungal; D008727:Methotrexate / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009693:Nucleic Acid Hybridization; D016366:Open Reading Frames / Q000235:genetics; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics
D003062:Codon; D001542:Benomyl; D008727:Methotrexate