Endosomal and AP-3-Dependent Vacuolar Trafficking Routes Make Additive Contributions to Candida albicans Hyphal Growth and Pathogenesis

Abstract

ABSTRACT Candida albicans mutants deficient in vacuolar biogenesis are defective in polarized hyphal growth and virulence. However, the specific vacuolar trafficking routes required for hyphal growth and virulence are unknown. In Saccharomyces cerevisiae , two trafficking routes deliver material from the Golgi apparatus to the vacuole. One occurs via the late endosome and is dependent upon Vps21p, while the second bypasses the endosome and requires the AP-3 complex, including Aps3p. To determine the significance of these pathways in C. albicans hyphal growth and virulence, aps3 Δ/Δ, vps21 Δ/Δ, and aps3 Δ/ Δ vps21 Δ/Δ mutant strains were constructed. Analysis of vacuolar morphology and localization of the vacuolar protein Mlt1p suggests that C. albicans Aps3p and Vps21p mediate two distinct transport pathways. The vps21 Δ/Δ mutant has a minor reduction in hyphal elongation, while the aps3 Δ/Δ mutant has no defect in hyphal growth. Interestingly, the aps3 Δ/ Δ vps21 Δ/Δ double mutant has dramatically reduced hyphal growth. Overexpression of the Ume6p transcriptional activator resulted in constitutive hyphal growth of wild-type, aps3 Δ/Δ, and vps21 Δ/Δ strains and formation of highly vacuolated subapical compartments. Thus, Ume6p-dependent transcriptional responses are sufficient to induce subapical vacuolation. However, the aps3 Δ/ Δ vps21 Δ/Δ mutant formed mainly pseudohyphae that lacked vacuolated compartments. The aps3 Δ/Δ strain was virulent in a mouse model of disseminated infection; the vps21 Δ/Δ mutant failed to kill mice but persisted within kidney tissue, while the double mutant was avirulent and cleared from the kidneys. These results suggest that while the AP-3 pathway alone has little impact on hyphal growth or virulence, it is much more significant when endosomal trafficking is disrupted.