Unusual Susceptibility of a Multidrug-Resistant Yeast Strain to Peptidic Antifungals

Abstract

The susceptibility of Saccharomyces cerevisiae JG436 multidrug transporter deletion mutant, Deltapdr5, to several antifungal agents was compared to that of JG436-derived JGCDR1 and JGCaMDR1 transformants, harboring the CDR1 and CaMDR1 genes, encoding the main drug-extruding membrane proteins of Candida albicans. The JGCDR1 and JGCaMDR1 yeasts demonstrated markedly diminished susceptibility to the azole antifungals, terbinafine and cycloheximide, while that to amphotericin B was unchanged. Surprisingly, JGCDR1 but not JGCaMDR1 cells showed enhanced susceptibility to peptidic antifungals, rationally designed compounds containing inhibitors of glucosamine-6-phosphate synthase. It was found that these antifungal oligopeptides, as well as model oligopeptides built of proteinogenic amino acids, were not effluxed from JGCDR1 cells. Moreover, they were taken up by these cells at rates two to three times higher than by JG436. The tested oligopeptides were rapidly cleaved to constitutive amino acids by cytoplasmic peptidases. Studies on the mechanism of the observed phenomenon suggested that an additive proton motive force generated by Cdr1p stimulated uptake of oligopeptides into JGCDR1 cells, thus giving rise to the higher antifungal activity of FMDP [N(3)-(4-methoxyfumaroyl)-L-2,3-diaminopropanoic acid]-peptides.

A new twist in ABC transporter mediated multidrug resistance – Pdr5 is a drug/proton co-transporter

“…Moreover, drug/proton co-transport would temporarily significantly increase the membrane potential of the plasma membrane thus effecting the ion and solute homeostasis and the membrane morphology of yeast cells (45-47). This compares well with results that were obtained previously for the Pdr5 homologue Cdr1 and MDR1 in vivo as it has been speculated that MDR1 as well as Cdr1 may transport protons from the intra-to the extracellular space and thus alkalize the cytoplasm (8, 38). However, the complexity of the cell environment of these in vivo experiments did not allow to unambiguously pinpoint observations to a single protein level nor for the experimenter to control all relevant critical cellular parameters.…”

Enzymes of UDP‐GlcNAc biosynthesis in yeast

“…Nva‐FMDP and Lys‐Nva‐FMDP, demonstrated strong chemotherapeutic activity in the murine model of disseminated histoplasmosis60 and candidiasis, as well as very little, if any, acute animal toxicity 55. Especially noteworthy is a remarkably high activity of FMDP‐peptides against fungal cells expressing the drug‐effluxing, ABC‐type membrane proteins 61. All these results obtained so far are sufficiently encouraging to consider GlcN‐6‐P synthase a really promising target for antifungal chemotherapy.…”

Inhibitors of the Candida albicans Major Facilitator Superfamily Transporter Mdr1p Responsible for Fluconazole Resistance

“…Hygromycin B has greater toxicity for cells over-expressing the ABC transporter CaCdr1p than the host strain (compare the zone of growth inhibition on Fig 5A with that on Fig 5C ). This is due to the electrogenic nature of CaCdr1p; the ATP hydrolysis it catalyses generates protons and the substrates it transports are partially positively charged [ 37 ]. The growth inhibitory zone for the AD/CaCDR1 strain exposed to 200 nmol hygromycin B was significantly larger than for AD/pABC3 and its size was not modified by placing adjacent disks containing up to 25 nmol compound A ( Fig 5D ).…”