Paradoxical Th1 activation and CTLA-4 regulation is beneficial during latent cryptococcosis

ABSTRACT Cryptococcus neoformans is the predominant causative agent of cryptococcal meningitis in immunocompromised individuals. Conversely, in immunocompetent individuals, C. neoformans establishes a latent pulmonary infection characterized by a paucity of clinical symptoms. Using a mouse inhalation model of latent C. neoformans infection, we previously showed that CD4 T cells are necessary for preventing fungal proliferation in the lungs. In the current study, we performed single-cell RNA sequencing (scRNA-seq) and found that the CD4 T-cell response was both highly heterogeneous and dichotomous during pulmonary C. neoformans infection, with concomitant expression of genes related to Th1 polarization ( Tbx21 and Ifng ) and immune regulation ( Ctla4 ). First, we demonstrated that cells with Th1-like phenotypes are necessary and sufficient to control latent infection via adoptive transfer of Tbet-positive cells into infection-matched CD4-depleted recipient mice. Second, scRNA-seq analysis revealed that the subpopulation of effector CD4 T cells that co-expressed Ctla4 and Gata3 was significantly higher than a subpopulation that co-expressed Ctla4 and Tbx21 . Furthermore, our data suggested that CTLA-4 upregulation is beneficial against C. neoformans infection as CTLA-4 blockade promoted fungal proliferation. Thus, we propose a model wherein Th1 control of latent C. neoformans infection is supported by CTLA-4 suppression of detrimental Th2 activation. IMPORTANCE CD4 T cells are a critical part of the adaptive immune response in many infections. Upon inhalation of the fungal pathogen Cryptococcus neoformans , CD4 T cells are necessary to control the initial latent infection and prevent disease. In immunocompromised individuals that lack CD4 T cells, the initial infection in the lungs can spread to the brain and cause lethal meningitis. In this study, we show that the CD4 T cells present during latent C. neoformans infection are heterogeneous with expression of diverse polarization and immune regulation markers that function in concert to prevent disease. Our studies highlight the previously unappreciated diversity and complex regulation of the CD4 T-cell response that is required to prevent disease in this important fungal pathogen.