Lipid Transport by Candida albicans Dnf2 Is Required for Hyphal Growth and Virulence

Candida albicans is a common cause of human mucosal yeast infections, and invasive candidiasis can be fatal. Antifungal medications are limited, but those targeting the pathogen cell wall or plasma membrane have been effective. Therefore, virulence factors controlling membrane biogenesis are potential targets for drug development. P4-ATPases contribute to membrane biogenesis by selecting and transporting specific lipids from the extracellular leaflet to the cytoplasmic leaflet of the bilayer to generate lipid asymmetry. A subset of heterodimeric P4-ATPases, including Dnf1-Lem3 and Dnf2-Lem3 from Saccharomyces cerevisiae, transport phosphatidylcholine (PC), phosphatidylethanolamine (PE), and the sphingolipid glucosylceramide (GIcCer). GIcCer is a critical lipid for Candida albicans polarized growth and virulence, but the role of GIcCer transporters in virulence has not been explored. Here, we show that the Candida albicans Dnf2 (CaDnf2) requires association with CaLem3 to form a functional transporter and flip fluorescent derivatives of GIcCer, PC, and PE across the plasma membrane. Mutation of conserved substrate-selective residues in the membrane domain strongly abrogates GIcCer transport and partially disrupts PC transport by CaDnf2. Candida strains harboring dnf2-null alleles (dnf2 Delta Delta) or point mutations that disrupt substrate recognition exhibit defects in yeast-to-hypha growth transition, filamentous growth, and virulence in systemically infected mice. The influence of CaONF1 deletion on the morphological phenotypes is negligible, although the dnf1 Delta Delta dnf2 Delta Delta strain was less virulent than the dnf2 Delta Delta strain. These results indicate that the transport of GIcCer and/or PC by plasma membrane P4-ATPases is important for the pathogenicity of Candida albicans.