Drug Susceptibilities of Yeast Cells Are Affected by Membrane Lipid Composition

Abstract

In the present study we have exploited isogenic erg mutants of Saccharomyces cerevisiae to examine the contribution of an altered lipid environment on drug susceptibilities of yeast cells. It is observed that erg mutants, which possess high levels of membrane fluidity, were hypersensitive to the drugs tested, i.e., cycloheximide (CYH), o-phenanthroline, sulfomethuron methyl, 4-nitroquinoline oxide, and methotrexate. Most of the erg mutants except mutant erg4 were, however, resistant to fluconazole (FLC). By using the fluorophore rhodamine-6G and radiolabeled FLC to monitor the passive diffusion, it was observed that erg mutant cells elicited enhanced diffusion. The addition of a membrane fluidizer, benzyl alcohol (BA), to S. cerevisiae wild-type cells led to enhanced membrane fluidity. However, a 10 to 12% increase in BA-induced membrane fluidity did not alter the drug susceptibilities of the S. cerevisiae wild-type cells. The enhanced diffusion observed in erg mutants did not seem to be solely responsible for the observed hypersensitivity of erg mutants. In order to ascertain the functioning of drug extrusion pumps encoding the genes CDR1 (ATP-binding cassette family) and CaMDR1 (MFS family) of Candida albicans in a different lipid environment, they were independently expressed in an S. cerevisiae erg mutant background. While the fold change in drug resistance mediated by CaMDR1 remained the same or increased in erg mutants, susceptibility to FLC and CYH mediated by CDR1 was increased (decrease in fold resistance). Our results demonstrate that between the two drug extrusion pumps, Cdr1p appeared to be more adversely affected by the fluctuations in the membrane lipid environment (particularly to ergosterol). By using 6-[(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino-hexanoyl] sphingosyl phosphocholine (a fluorescent analogue of sphingomyelin), a close interaction between membrane ergosterol and sphingomyelin which appears to be disrupted in erg mutants is demonstrated. Taken together it appears that multidrug resistance in yeast is closely linked to the status of membrane lipids, wherein the overall drug susceptibility phenotype of a cell appears to be an interplay among drug diffusion, extrusion pumps, and the membrane lipid environment.

Influences of cinnamic aldehydes on H+ extrusion activity and ultrastructure of Candida

“…Five microlitres of fivefold diluted yeast culture was spotted onto YNB plates in the absence (control) and in the presence of different concentrations of the test compounds. Growth differences were recorded following incubation of the plates for 48 h at 30 uC (Mukhopadhyay et al, 2002).…”

Fcr1p Inhibits Development of Fluconazole Resistance in Candida albicans by Abolishing CDR1 Induction

“…30) Approximately 10 7 cells from an overnight culture were incubated in 200 ml of YPD medium and grown for 5 h at 30°C. The cells were pelleted and washed three times with phosphate-buffered saline (PBS) buffer without glucose.…”

Decrease of H2O2 Plasma Membrane Permeability during Adaptation to H2O2 in Saccharomyces cerevisiae

“…Further supporting the importance of the plasma membrane is the observation that after a nonlethal dose of H 2 O 2 , the levels of mRNA that are more repressed are those corresponding to the enzyme HMG-CoA reductase (8). This is the rate-limiting enzyme of the synthesis of sterols, which are key components of the plasma membrane controlling its fluidity (9) and consequently the permeability of this membrane to a species like H 2 O 2 (10).…”

“…The two strains used (erg3⌬ and erg6⌬) have a defect in the pathway of ergosterol biosynthesis: erg3⌬ mutants lack a C-5 desaturase producing ergosta-7,22-dienol instead of ergosterol (31), and erg6⌬ mutants lack a C-24 methyltransferase producing zymosterol and colesta-5,7,24-trienol instead of ergosterol (32). In both mutants the membrane biophysical properties are changed, resulting in increased permeability to lipophilic compounds (9). As can be observed in Table III, these mutants show a higher H 2 O 2 consumption rate constant when compared with wild type cells, despite a similar cytochrome c peroxidase activity and similar or lower catalase activity in permeabilized cells.…”