Sporothrix globosa melanin regulates autophagy via the TLR2 signaling pathway in THP-1 macrophages

Author summaryMelanin as an important virulence factor for Sporothrix globosa, analysis of the effects on host immunity, is of great significance for guidance regarding future directions of studies on the mechanism underling fungal pathogenesis. In recent years, accumulating evidence supports a role for autophagy as a host defense mechanism to counteract the evasion strategies of viral, bacterial and parasitic pathogens. Our study aimed to investigate the effect of melanin on autophagy in macrophages, with particular attention in Sporothrix spp. to explore the mechanism of melanin interaction with Toll-like receptor (TLR)-induced pathways. In addition, this work also served as an important theoretical supplement for our previous research on S. globosa melanin. We believe that our research makes a significant contribution to the melanin pigment in fungi because although a lot of progress is being made in the field of pathogenic fungi, most studies are in animal macrophages in vitro studies; exploring the macrophage autophagy will provide a novel immune defense ability of S. globosa melanin to inhibit macrophage. Melanin, an important virulence factor of pathogenic fungi, has been shown to suppress host immune responses in multiple ways. Autophagy is a vital cellular mechanism underlying the host's innate immunity against microbial infections. However, the potential influence of melanin on autophagy has not been explored. We investigated the effect of melanin on autophagy in macrophages, which play a key role in controlling Sporothrix spp. infection, as well as the mechanism of melanin interaction with Toll-like receptor (TLR)-induced pathways. Sporothrix globosa conidia (wild-type and melanin-deficient mutant strains) or yeast cells were co-cultured with THP-1 macrophages to demonstrate that, although S. globosa infection led to the activation of autophagy-related proteins and increased autophagic flux, S. globosa melanin suppressed macrophage autophagy. Incubation with S. globosa conidia also increased the expression levels of reactive oxygen species and multiple proinflammatory cytokines (interleukin-6, tumor necrosis factor-alpha, interleukin-1 beta and interferon-gamma) in macrophages. These effects were attenuated as melanin presented. Furthermore, while S. globosa conidia significantly increased the expression of both TLR2 and TLR4 in macrophages, the knockdown of TLR2, but not TLR4, with small interfering RNA suppressed autophagy. Overall, this study revealed the novel immune defense ability of S. globosa melanin to inhibit macrophage functionality by resisting macrophage autophagy through the regulation of TLR2 expression.