Abstract
Candidalysin is a cytolytic peptide produced by the opportunistic fungal pathogen Candida albicans. This peptide is a key virulence factor in mouse models of mucosal and hematogenously disseminated candidiasis. Despite intense interest in the role of candidalysin in C. albicans pathogenicity, its host cell targets have remained elusive. To fill this knowledge gap, we performed a genome-wide loss-of-function CRISPR screen in a human oral epithelial cell line to identify specific host factors required for susceptibility to candidalysin-induced cellular damage. Among the top hits were XYLT2, B3GALT6 and B3GAT3, genes that function in glycosaminoglycan (GAG) biosynthesis. Deletion of these genes led to the absence of GAGs such as heparan sulfate on the epithelial cell surface and increased resistance to damage induced by both candidalysin and live C. albicans. Biophysical analyses including surface plasmon resonance and atomic force and electron microscopy indicated that candidalysin physically binds to sulfated GAGs, facilitating its oligomerization or enrichment on the host cell surface. The addition of exogenous sulfated GAGs or the GAG analogue dextran sulfate protected cells against candidalysin-induced damage. Dextran sulfate, but not non-sulfated dextran, also inhibited epithelial cell endocytosis of C. albicans and fungal-induced epithelial cell cytokine and chemokine production. In a murine model of vulvovaginal candidiasis, topical dextran sulfate administration reduced host tissue damage and decreased intravaginal IL-1β and neutrophil levels. Collectively, these data indicate that GAGs are epithelial cell targets of candidalysin and can be used therapeutically to protect cells from candidalysin-induced damage.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
J.L. and S.F. initiated and designed the project. S.F secured funding for the project. J.L. carried out the CRISPR-Cas9 screen. J.L. constructed the cell knockouts. M.Y. and S.F. provided candidalysin and J.L. performed the candidalysin damage assays, immunofluorescence imaging, and immune-blots. T.P. and J.L. did the fungal invasion assays. H.L. and J.L. performed the IL-1β, GM-CSF and CXCL8 ELISAs, flow cytometry and calcium flux assays. M.T, and J.L. did the LDH assays. K.G. and G.K. performed the electron microscopy and atomic force microscopy imaging. F.Z., J.D. and R.L. performed the SPR assays and acquisition of funding. C.R., R.P. and F.B. did the dye release and C-laurdan assays. B.P., J.M. and N.S. carried out the animal infection experiment and related assays. J.L. and S.F. wrote the manuscript with input, revision and review from all co-authors.
Data availability
The original sequencing data were deposited to the NCBI under BioProject PRJNA1081917. All data from this study are presented in the manuscript and supplementary figures and tables.
