FIG 2 .
The rim101Δ mutant cell wall induces TNF-α secretion from BMMs in a contact-dependent and capsule-independent manner. (A) WT and rim101Δ cells were coincubated with BMMs at an MOI of 10:1 C. neoformans/BMM cells, either “in direct contact” or “separated” by a Transwell system. (n = 3 for each strain/condition.) (B) BMM TNF-α levels (in picograms per milliliter) after 6 h of incubation with 10 µg/ml purified cell wall material from the indicated fungal strains. Data represent means of results from three replicate samples for three independent cell wall preparations (n = 9 for each strain). (C) BMMs were coincubated for 6 h with UV-killed fungal cells that had been precultured in either YPD (rich) medium or tissue culture (TC), CO2-independent medium. TNF-α levels in the culture medium were normalized to a control incubation with no added fungal cells. Primary data for TNF-α levels are listed in Table S2. Data represent means of results from three replicate samples for two independent experiments (n = 6 for each strain/condition). (D) BMMs were coincubated with the indicated strains for 6 h. The gray line separates strains normalized by cell number (WT and rim101Δ strains) and by weight (cap59Δ strain or cap59Δ rim101Δ strain). Data represent means of results from three replicate samples (n = 9). Two-way (A) and one-way (B, C, and D) ANOVA and Tukey’s multiple-comparison test were used to compare means. ***, P < 0.001; **, P < 0.01; *, P < 0.05. Error bars represent standard errors of the means.