A permease encoded by STL1 is required for active glycerol uptake by Candida albicans

Abstract

Candida albicansaccumulates large amounts of the polyols glycerol andd-arabitol when the cells are exposed to physiological conditions relevant to stress and virulence in animals. Intracellular concentrations of glycerol are determined by rates of glycerol production and catabolism and of glycerol uptake and efflux through the plasma membrane. We and others have studied glycerol production inC. albicans, but glycerol uptake byC. albicanshas not been studied. In the present study, we found that [14C]glycerol uptake byC. albicansSC5314 was (i) accumulative; (ii) dependent on proton-motive force; (iii) unaffected by carbon source; and (iv) unaffected by large molar excesses ofd-arabitol or other polyols. The respectiveKmandVmaxvalues were 2.1 mM and 460 μmol h−1(g dry wt)−1in glucose medium and 2.6 mM and 268 μmol h−1(g dry wt)−1in glycerol medium. To identify theC. albicansglycerol uptake protein(s), we cloned theC. albicanshomologues of theSaccharomyces cerevisiaegenesGUP1andSTL1, both of which are known to be involved in glycerol transport. When multicopy plasmids encodingC. albicans STL1,C. albicans STL2andC. albicans GUP1were introduced into the correspondingS. cerevisiaenull mutants, the transformants all acquired the ability to grow on minimal glycerol medium; however, onlyS. cerevisiae stl1null mutants transformed withC. albicans STL1actively took up extracellular [14C]glycerol. When both chromosomal alleles ofC. albicans STL1were deleted fromC. albicansBWP17, the resultingstl1null mutants grew poorly on minimal glycerol medium, and their ability to transport [14C]glycerol into the cell was markedly reduced. In contrast, deletion of both chromosomal alleles ofC. albicans STL2or ofC. albicans GUP1had no significant effects on [14C]glycerol uptake or the ability to grow on minimal glycerol medium. Northern blot analysis indicated thatC. albicans STL1was expressed in both glucose and glycerol media, conditions under which we detected wild-type active glycerol uptake. Furthermore,STL1was highly expressed in salt-stressed cells; however, thestl1null mutant was no more sensitive to salt stress than wild-type controls. We also detected high levels ofSTL2expression in glycerol-grown cells, even though deletion of this gene did not influence glycerol uptake activity in glycerol-grown cells. We conclude from the results above that a plasma-membrane H+symporter encoded byC. albicans STL1actively transports glycerol intoC. albicanscells.