Rivera et al. (2025). Infection and Immunity. doi: 10.1128/iai.00355-25
…at 37°C in activating medium (DME medium, 50 U/mL of IFN- γ, 1 g/mL of lipopolysaccharide [LPS]). Phosphate-buffered saline pH 7.4 (PBS)-washed fungal cells were opsonized with 20% mouse complement (Pel-Freez, AK) and added to the macrophages at an effector-to-target ratio of 1:2. Phagocytosis was allowed to occur for 2 h at 37°C in 10% CO 2 . The cells were then washed three times with 1X PBS and fixed with…
Pel-Freez products cited: Mouse Complement (35025)
Abstract
ABSTRACT Currently, no fungal vaccine exists for clinical use, while fungal infections are responsible for over 1.5 million deaths every year. Our previous studies identified a Cryptococcus neoformans mutant strain fbp1 Δ as a potential vaccine candidate. This strain contains deletion of the F-box protein Fbp1, a key subunit of the SCF E3 ligase complex necessary for ubiquitin-mediated proteolysis. Vaccination with heat-killed fbp1 Δ (HK- fbp1 ) can elicit an interferon gamma (IFN-γ)-dependent Type 1 immune response and provide protection against C. neoformans and its sibling species C. gattii . However, we have yet to decipher the immunogenic factor(s) expressed by the fbp1 ∆ mutant that are responsible for the induction of the protective immune response. In this study, we have identified that the capsule plays an important role in HK- fbp1 vaccine-mediated protection as acapsular HK- fbp1 cells showed diminished protection against wild-type challenge. Additionally, our studies have shown that Cytokine Inducing Glycoprotein 1 (Cig1), a GPI-anchored mannoprotein, is regulated by Fbp1 and contributes to the immunogenicity of HK- fbp1 . Deletion of Cig1 in the fbp1 Δ background resulted in decreased recruitment of antifungal effector T cells and diminished production of protective inflammatory cytokines by the host. Furthermore, loss of Cig1 in the fbp1 Δ mutant resulted in reduced protection in vaccination survival studies at lower vaccine inoculum doses compared to HK- fbp1 . In aggregate, these findings demonstrate Cig1 is an antigen contributing to the immunogenicity of HK- fbp1 that may be utilized to further optimize the HK- fbp1 fungal vaccine as a tool in the arsenal against invasive fungal infections.