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Inflammasome-mediated GSDMD activation facilitates escape of Candida albicans from macrophages.

ABSTRACT: Candida albicans is the most common cause of fungal sepsis. Inhibition of inflammasome activity confers resistance to polymicrobial and LPS-induced sepsis; however, inflammasome signaling appears to protect against C. albicans infection, so inflammasome inhibitors are not clinically useful for candidiasis. Here we show disruption of GSDMD, a known inflammasome target and key pyroptotic cell death mediator, paradoxically alleviates candidiasis, improving outcomes and survival of Candida-infected mice. Mechanistically, C. albicans hijacked the canonical inflammasome-GSDMD axis-mediated pyroptosis to promote their escape from macrophages, deploying hyphae and candidalysin, a pore-forming toxin expressed by hyphae. GSDMD inhibition alleviated candidiasis by preventing C. albicans escape from macrophages while maintaining inflammasome-dependent but GSDMD-independent IL-1β production for anti-fungal host defenses. This study demonstrates key functions for GSDMD in Candida's escape from host immunity in vitro and in vivo and suggests that GSDMD may be a potential therapeutic target in C. albicans-induced sepsis.

SUBMITTER: Ding X

PROVIDER: S-EPMC8602704 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The fungal pathogen Candida albicans causes macrophage death and escapes, but the molecular mechanisms remained unknown. Here we used live-cell imaging to monitor the interaction of C. albicans with macrophages and show that C. albicans kills macrophages in two temporally and mechanistically distinct phases. Early upon phagocytosis, C. albicans triggers pyroptosis, a proinflammatory macrophage death. Pyroptosis is controlled by the developmental yeast-to-hypha transition of Candida. When pyroptosis is inactivated, wild-type C. albicans hyphae cause significantly less macrophage killing for up to 8 h postphagocytosis. After the first 8 h, a second macrophage-killing phase is initiated. This second phase depends on robust hyphal formation but is mechanistically distinct from pyroptosis. The transcriptional regulator Mediator is necessary for morphogenesis of C. albicans in macrophages and the establishment of the wild-type surface architecture of hyphae that together mediate activation of macrophage cell death. Our data suggest that the defects of the Mediator mutants in causing macrophage death are caused, at least in part, by reduced activation of pyroptosis. A Mediator mutant that forms hyphae of apparently wild-type morphology but is defective in triggering early macrophage death shows a breakdown of cell surface architecture and reduced exposed 1,3 ?-glucan in hyphae. Our report shows how Candida uses host and pathogen pathways for macrophage killing. The current model of mechanical piercing of macrophages by C. albicans hyphae should be revised to include activation of pyroptosis by hyphae as an important mechanism mediating macrophage cell death upon C. albicans infection. IMPORTANCE Upon phagocytosis by macrophages, Candida albicans can transition to the hyphal form, which causes macrophage death and enables fungal escape. The current model is that the highly polarized growth of hyphae results in macrophage piercing. This model is challenged by recent reports of C. albicans mutants that form hyphae of wild-type morphology but are defective in killing macrophages. We show that C. albicans causes macrophage cell death by at least two mechanisms. Phase 1 killing (first 6 to 8 h) depends on the activation of the pyroptotic programmed host cell death by fungal hyphae. Phase 2 (up to 24 h) is rapid and depends on robust hyphal formation but is independent of pyroptosis. Our data provide a new model for how the interplay between fungal morphogenesis and activation of a host cell death pathway mediates macrophage killing by C. albicans hyphae.

Project description:Candida albicans cells depend on the energy derived from amino acid catabolism to induce and sustain hyphal growth inside phagosomes of engulfing macrophages. The concomitant deamination of amino acids is thought to neutralize the acidic microenvironment of phagosomes, a presumed requisite for survival and initiation of hyphal growth. Here, in contrast to an existing model, we show that mitochondrial-localized NAD+-dependent glutamate dehydrogenase (GDH2) catalyzing the deamination of glutamate to α-ketoglutarate, and not the cytosolic urea amidolyase (DUR1,2), accounts for the observed alkalization of media when amino acids are the sole sources of carbon and nitrogen. C. albicans strains lacking GDH2 (gdh2-/-) are viable and do not extrude ammonia on amino acid-based media. Environmental alkalization does not occur under conditions of high glucose (2%), a finding attributable to glucose-repression of GDH2 expression and mitochondrial function. Consistently, inhibition of oxidative phosphorylation or mitochondrial translation by antimycin A or chloramphenicol, respectively, prevents alkalization. GDH2 expression and mitochondrial function are derepressed as glucose levels are lowered from 2% (~110 mM) to 0.2% (~11 mM), or when glycerol is used as primary carbon source. Using time-lapse microscopy, we document that gdh2-/- cells survive, filament and escape from primary murine macrophages at rates indistinguishable from wildtype. In intact hosts, such as in fly and murine models of systemic candidiasis, gdh2-/- mutants are as virulent as wildtype. Thus, although Gdh2 has a critical role in central nitrogen metabolism, Gdh2-catalyzed deamination of glutamate is surprisingly dispensable for escape from macrophages and virulence. Consistently, using the pH-sensitive dye (pHrodo), we observed no significant difference between wildtype and gdh2-/- mutants in phagosomal pH modulation. Following engulfment of fungal cells, the phagosomal compartment is rapidly acidified and hyphal growth initiates and sustained under consistently acidic conditions within phagosomes. Together, our results demonstrate that amino acid-dependent alkalization is not essential for hyphal growth, survival in macrophages and hosts. An accurate understanding of the microenvironment within macrophage phagosomes and the metabolic events underlying the survival of phagocytized C. albicans cells and their escape are critical to understanding the host-pathogen interactions that ultimately determine the pathogenic outcome.

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Inflammasome-mediated GSDMD activation facilitates escape of Candida albicans from macrophages.

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