Probing the role of point mutations in the cyp51A gene from Aspergillus fumigatus in the model yeast Saccharomyces cerevisiae.
Azole-resistant strains of Aspergillus fumigatus have been detected and the underlying molecular mechanisms of resistance characterized. Point mutations in the cyp51A gene have been proved to be related to azole resistance in A. fumigatus clinical strains and with different resistance profiles depending on the amino acid change (G54E, G54V, G54R, G54W, M220V, M220K, M220T, M220I). The aim of this work was to express A. fumigatus cyp51A genes in the yeast Saccharomyces cerevisiae in order to better assess the contribution of each independent amino acid substitution to resistance. A tetracycline regulatable system allowing repression of the endogenous essential ERG11 gene was used. The expression of Aspergillus cyp51A alleles could efficiently restore the absence of ERG11 in S. cerevisiae. In general, S. cerevisiae clones expressing. A. fumigatus cyp51A alleles from azole-resistant isolates showed higher MICs to all azoles tested than those expressing alleles from susceptible isolates. The azole susceptibility profiles obtained in S. cerevisiae upon expression of specific cyp51A alleles recapitulated susceptibility profiles observed from their A. fumigatus origins. In conclusion this work supports the concept that characteristics of specific A. fumigatus cyp51A alleles could be investigated in the heterologous host S. cerevisiae.
Alcazar-Fuoli
L
L
Mycology Reference Laboratory, Centro Nacional de Microbiologia, Instituto de Salud Carlos III, Majadahonda, Madrid, Spain.
Mellado
E
E
Cuenca-Estrella
M
M
Sanglard
D
D
D019943:Amino Acid Substitution / Q000235:genetics; D000935:Antifungal Agents / Q000494:pharmacology*; D001232:Aspergillus fumigatus / Q000187:drug effects*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004252:DNA Mutational Analysis; D025141:Drug Resistance, Fungal; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D008826:Microbial Sensitivity Tests; D020125:Mutation, Missense; D017354:Point Mutation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics
D000935:Antifungal Agents; D005656:Fungal Proteins; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; C465526:cytochrome P-450 CYP51A, Aspergillus