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

¹ Department of Internal Medicine, Section of Infectious Diseases, Yale University and VA Connecticut Healthcare System, 950 Campbell Avenue (111-I), West Haven, CT 06516, USA
² Centro de Biologia Molecular e Ambiental (CBMA), Departamento de Biologia/Universidade do Minho, Campus de Gualtar, 4710-057, Braga, Portugal
³ Division of Infectious Diseases, Department of Medicine, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, NRC-3, Portland, OR 97239, USA

Candida albicans accumulates large amounts of the polyols glycerol and D-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 in C. albicans, but glycerol uptake by C. albicans has not been studied. In the present study, we found that [¹⁴C]glycerol uptake by C. albicans SC5314 was (i) accumulative; (ii) dependent on proton-motive force; (iii) unaffected by carbon source; and (iv) unaffected by large molar excesses of D-arabitol or other polyols. The respective K_m and V_max values were 2.1 mM and 460 µmol h^–1 (g dry wt)^–1 in glucose medium and 2.6 mM and 268 µmol h^–1 (g dry wt)^–1 in glycerol medium. To identify the C. albicans glycerol uptake protein(s), we cloned the C. albicans homologues of the Saccharomyces cerevisiae genes GUP1 and STL1, both of which are known to be involved in glycerol transport. When multicopy plasmids encoding C. albicans STL1, C. albicans STL2 and C. albicans GUP1 were introduced into the corresponding S. cerevisiae null mutants, the transformants all acquired the ability to grow on minimal glycerol medium; however, only S. cerevisiae stl1 null mutants transformed with C. albicans STL1 actively took up extracellular [¹⁴C]glycerol. When both chromosomal alleles of C. albicans STL1 were deleted from C. albicans BWP17, the resulting stl1 null 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 of C. albicans STL2 or of C. albicans GUP1 had no significant effects on [¹⁴C]glycerol uptake or the ability to grow on minimal glycerol medium. Northern blot analysis indicated that C. albicans STL1 was expressed in both glucose and glycerol media, conditions under which we detected wild-type active glycerol uptake. Furthermore, STL1 was highly expressed in salt-stressed cells; however, the stl1 null mutant was no more sensitive to salt stress than wild-type controls. We also detected high levels of STL2 expression 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 by C. albicans STL1 actively transports glycerol into C. albicans cells.

Correspondence
Gerald Kayingo
gerald.kayingo{at}yale.edu

A supplementary table of primers is available with the online version of this paper.