Nanoscopic Cell Wall Architecture of an Immunogenic Ligand in Candida Albicans during Antifungal Drug Treatment

Candida albicans is a common cause of high-mortality disseminated fungal infections. Morphotype switching between yeasts and filamentous hyphae is important for virulence. Innate immune cells use C-type lectins to recognize Candida cell wall polysaccharides. In particular, Dectin-1 recognizes fungal cell wall beta-glucan, leading to phagocytosis and pro-inflammatory responses. However, C. albicans partially evades innate immune recognition by restricting glucan surface exposure. Treatment with the antifungal caspofungin increases surface exposure of beta-glucan and leukocyte phagocytosis. Our objective was to better understand the nanoscale structure of glucan exposures that C. albicans presents to leukocytes. We probed exposed glucan with AF647-soluble Dectin-1 ectodomain and used direct Stochastic Optical Reconstruction Microscopy (dSTORM) to quantify glucan exposure on C. albicans yeast and hyphal cell walls, both in the resting state and after treatment with caspofungin. Strikingly, the vast majority of glucan exposure sites were consistent with single receptor binding sites in all cases. In resting yeasts, the multiple-probe exposure sites were few and small. After caspofungin treatment, multi-probe exposures were larger and surface density of glucan exposures was increased. Untreated C. albicans hyphae had a higher glucan exposure surface density than yeasts. Lateral hyphal cell walls imaged by dSTORM underwent little quantitative or qualitative change in glucan exposure geometry after drug treatment, but confocal imaging revealed higher glucan exposure after caspofungin treatment at the actively growing and remodeling cell walls of hyphal tips. Similarly, in budding yeasts, we found consistently higher glucan exposure in the actively remodeling cell wall of daughter cells. These findings enable future mechanistic investigation of the contribution of glucan nanopatterning to C. albicans evasion of innate immune recognition via Dectin-1 and the use of nanopatterning of glucan for therapeutic interventions in fungal infectious disease.