Tumor Necrosis Factor Receptor–Associated Factor 1 Inhibits Macrophage Killing of Sporothrix schenckii by Enhancing Expression and Activity of Inducible Nitric Oxide Synthase

BACKGROUND: Sporotrichosis is a chronic, deep fungal infection of skin caused by Sporothrix schenckii. Macrophages are predominant in S. schenckii-infected skin and are able to phagocytize and kill the fungus. Local hyperthermia is effective for treating sporotrichosis, but its mechanism of action is still not fully understood. METHODS: Using single-cell RNA sequencing of sporotrichosis lesions, coupled with in vitro and in vivo sporotrichosis models, we investigated the role of tumor necrosis factor receptor-associated factor 1 (TRAF1) and nitric oxide synthase, inducible (NOS2). Mechanistic studies included coimmunoprecipitation, ubiquitination assays, and site-directed mutagenesis. Therapeutic mechanisms of hyperthermia were evaluated in vivo and in vitro. RESULTS: We demonstrated for the first time that TRAF1 could delay the healing of sporotrichosis by inhibiting phagocytosis and killing of macrophages with S. schenckii. This effect of TRAF1 is caused by binding NOS2 to regulate its expression and enzymatic activity, through inhibition of NOS2 ubiquitination and subsequent proteasome-induced degradation. Our team's previous research has demonstrated the efficacy of hyperthermia in treating sporotrichosis. Our experiments indicate that hyperthermia can down-regulate the expression of TRAF1 and NOS2 in macrophages. CONCLUSIONS: We identify TRAF1-mediated stabilization of NOS2 as a key immune evasion mechanism in S. schenckii infection. Local hyperthermia represents a targeted therapy against this pathway, offering a novel strategy for enhancing the therapeutic effect of hyperthermia.