Defects in intracellular trafficking of fungal cell wall synthases lead to aberrant host immune recognition
Fig 5
Mar1 protein localization is dynamic over time in tissue culture medium.
(A) Localization of a Mar1-GFP fusion protein was assessed after cells were incubated for 16–18 hours in YPD (30°C) or TC (37°C). Live cells were imaged using DeltaVision deconvolution fluorescent microscopy with the GFP filter. Images were deconvolved using softWoRx software. Bar, 10 μM. (B) Cells were incubated and imaged as above. 3D-projections were generated from z-stacked images using ImageJ/Fiji software and are pseudo-colored to indicate brightness of staining from yellow to blue. Bar, 10 μM. (C) A Mar1-GFP expression strain was pre-incubated in YPD medium to mid-log phase and transferred to TC medium at time zero. Cells were incubated for the indicated time in TC at 37°C with shaking and imaged by fluorescent microscopy using the GFP filter. Bar, 10 μM. (D) Quantification of staining by FITC-conjugated WGA. The WT and mar1Δ strains were incubated to mid-log phase in YPD medium and transferred to TC medium at time zero. Exposed chitooligomers in the cell wall were stained with FITC-conjugated WGA, imaged by fluorescent microscopy using the GFP filter, and average fluorescence was quantified by ImageJ/Fiji software. ****, p < 0.0001 as determined by two-way ANOVA with Sidak’s multiple comparisons test.