MHC-I Upregulation and Increased Glucose Dependence Define Innate Antifungal Immune Responses to Cryptococcus neoformans.

BACKGROUND

Cryptococcus neoformans, the etiologic agent of cryptococcosis, can survive and replicate within host immune cells, contributing to development of cryptococcal meningitis. Within the lung, C neoformans interacts with innate immune subsets, including macrophages and dendritic cells (DCs). Our previous work demonstrated that Ly6c- monocyte-like macrophages restrict fungal proliferation, whereas CD11b+ DCs promote fungal growth. Transcriptomic profiling revealed differential expression of MHC-I gene H2-K1, calreticulin, and metabolic genes. Therefore, we hypothesized that H2-K1 and metabolic pathways modulate intracellular fungal fate within pulmonary macrophages and DCs.

METHODS

H2-K1 expression was knocked down by siRNA in J774 macrophages and bone marrow-derived DCs induced by granulocyte-macrophage colony-stimulating factor. Antifungal activity was determined by colony-forming unit enumeration. Phagocytic uptake, cathepsin B activity, Nos2 expression, and reactive oxygen species production were quantified by flow cytometry. Metabolic dependencies were evaluated through SCENITH assay (single-cell energetic metabolism by profiling translational inhibition).

RESULTS

H2-K1 knockdown in J774 cells significantly reduced antifungal activity as compared with controls. H2-K1 knockdown also decreased calreticulin gene expression, suggesting that H2-k1 has downstream effects in the pathway. H2-K1 silencing did not significantly alter phagocytic uptake, cathepsin B activity, or Nos2 expression. However, reactive oxygen species production was significantly reduced in H2-K1 knockdown J774 cells, indicating impaired responses. SCENITH analysis revealed that antifungal macrophages shift their metabolism toward glucose oxidation for ATP generation, which is important for antifungal activity.

CONCLUSIONS

These findings indicate that metabolic reprogramming is critical for effective antifungal responses. Our data provide a foundation for future studies aimed at targeting host immune and metabolic pathways to enhance antifungal immunity.