Candida glabrata is an increasingly important opportunistic human fungal pathogen which appears to be intrinsically resistant to the triazole antifungal fluconazole. The expression and drug efflux activity of the ATP binding cassette transporters Cdr1p and Pdh1p are thought to contribute to high level azole resistance in C. glabrata. We have evaluated the function of these transporters and their pumping characteristics through their heterologous hyper-expression in a Saccharomyces cerevisiae strain that is deleted in seven major drug efflux transporters in order to minimize the background drug efflux activity. Although both Cdr1p- and Pdh1p-expressing strains CDR1-AD and PDH1-AD acquired multiple resistances to structurally unrelated compounds, CDR1-AD showed, in most cases, higher levels of resistance than PDH1-AD. We found that exposure to fluconazole rapidly induced expression of a 169 kDa and a 61 kDa protein in plasma membrane fractions of C. glabrata cells. Mass spectrometric peptide fingerprinting identified the 169 kDa band as Cdr1p and the 61-kDa protein as the azole target lanosterol 14˜-demethylase (Erg11p). We also found that fermenting C. glabrata cells were less sensitive to azole antifungals than cells grown on glycerol or under aerobic conditions. Higher levels of both CDR1 and ERG11 but not PDH1 mRNA expression were detected in the fermenting C. glabrata cells. These results indicate that Cdr1p and Erg11p make a significant, rapidly inducible, contribution to the innate fluconazole resistance in C. glabrata that also depends on the metabolic status of the cells.
Full conference title:
The 15 th Congress of the International Society for Human and Animal Mycology
- ISHAM 15th (2003)