Fig. 3. ΔaceA strains accumulate more copper during macrophage encounters.
(A) Western blot against mouse Ctr1 and GAPDH of murine bone marrow derived macrophages activated with GM-CSF that were unchallenged or challenged with A. fumigatus spores for 2h.
(B) Immuno-staining against mouse ATP7A of murine bone marrow derived macrophages activated with GM-CSF that were unchallenged or challenged with A. fumigatus spores for 2h. Scale bars are 10 μM.
(C) Colony forming units (CFU) of fungal strains after incubation with murine bone marrow derived macrophages activated with GM-CSF for 2h. Experiments were carried out in triplicates; error bars represent standard deviations and statistical significance is indicated by p values.
(D) Total copper concentration of unchallenged 3 × 107 spores (solid) and 3 × 107 spores incubated with 1 × 107 GM-CSF activated bone marrow derived murine macrophages for 2h. Experiments were carried out in triplicates; error bars represent standard deviations and statistical significance is indicated by p values.
(E) Total copper concentration of 1 × 107 GM-CSF activated bone marrow derived murine macrophages incubated with 3 × 107 spores of the indicated A. fumigatus strains for 2h. Experiments were carried out in triplicates; error bars represent standard deviations and statistical significance is indicated by p values.
(F) Colony forming units (CFU) of fungal strains from whole zebrafish larvae at 24 hours post microinjection. Genetic inhibition of ATP7A was obtained with morpholino-mediated knockdown (ATP7AMO). Data shown are pooled from four independent experimental replicates where significance is indicated by p values as determined by a least squares means analysis.
(G) Colony forming units (CFU) of fungal strains after incubation with murine alveolar macrophages supplemented with or without 50 μM tetrathiomolybdate (TTM) for 2 h. Experiments were carried out in triplicates; error bars represent standard deviations and statistical significance is indicated by p values.