A fungal lytic polysaccharide monooxygenase is required for cell wall integrity, thermotolerance, and virulence of the fungal human pathogen Cryptococcus neoformans
Fig 5
CnCel1 affects melanization and capsule architecture.
(A) Melanization in the absence and presence of exogenous copper. Indicated strains were incubated on L-DOPA plates at 30°C or 37°C, with and without supplementation with exogenous CuSO4. Melanin formation was assessed at 1–2 days as indicated. (B-E) Surface capsule formation and architecture. The WT and cel1Δ mutant were incubated for 3 days in CIM tissue culture medium to induce capsule. Surface capsule characteristics were assessed by (B) India ink counter-staining and (C) SEM. Representative images are shown from 3 independent experiments. Quantification of capsule size (D) and cell body size (E) was performed using ImageJ/Fiji from static images of india ink-stained cells. Mean values from >100 cells per sample (+/- SEM) from 3 biological replicates are shown as bar graph. Data were plotted, and an unpaired t-test was performed using GraphPad Prism. (F) Electromobility of exopolysaccharide. The WT and cel1Δ mutant were incubated in CIM for 3 days. Exopolysaccharide in the culture supernatant was assessed for relative size and gel motility by agarose gel electrophoresis, transferred to a Nylon membrane, and probed with anti-GXM antibody 18B7. Representative image of multiple replicates is shown.