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

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, aps3Δ/Δ, vps21Δ/Δ, and aps3Δ/Δ vps21Δ/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 vps21Δ/Δ mutant has a minor reduction in hyphal elongation, while the aps3Δ/Δ mutant has no defect in hyphal growth. Interestingly, the aps3Δ/Δ vps21Δ/Δ double mutant has dramatically reduced hyphal growth. Overexpression of the Ume6p transcriptional activator resulted in constitutive hyphal growth of wild-type, aps3Δ/Δ, and vps21Δ/Δ strains and formation of highly vacuolated subapical compartments. Thus, Ume6p-dependent transcriptional responses are sufficient to induce subapical vacuolation. However, the aps3Δ/Δ vps21Δ/Δ mutant formed mainly pseudohyphae that lacked vacuolated compartments. The aps3Δ/Δ strain was virulent in a mouse model of disseminated infection; the vps21Δ/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.