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Author(s): Samarchith P Kurup [1]; Nyamekye Obeng-Adjei [2]; Scott M Anthony [1]; Boubacar Traore [3]; Ogobara K Doumbo [3]; Noah S Butler [1, 4]; Peter D Crompton [2]; John T [...]

Prostaglandin D2 (PGD2), an eicosanoid with both pro- and antiinflammatory properties, is the most abundantly expressed prostaglandin in the brain. Here we show that PGD2 signaling through the D-prostanoid receptor 1 (DP1) receptor is necessary for optimal microglia/macrophage activation and IFN expression after infection with a neurotropic coronavirus. Genome-wide expression analyses indicated that PGD2/DP1 signaling is required for upregulation of a putative inflammasome inhibitor, PYDC3, in CD11b+ cells in the CNS of infected mice. Our results also demonstrated that, in addition to PGD2/DP1 signaling, type 1 IFN (IFN-I) signaling is required for PYDC3 expression. In the absence of Pydc3 up-regulation, IL-1β expression and, subsequently, mortality were increased in infected DP1-/- mice. Notably, survival was enhanced by IL1 receptor blockade, indicating that the effects of the absence of DP1 signaling on clinical outcomes were mediated, at least in part, by inflammasomes. Using bone marrow-derived macrophages in vitro, we confirmed that PYDC3 expression is dependent upon DP1 signaling and that IFN priming is critical for PYDC3 up-regulation. In addition, Pydc3 silencing or overexpression augmented or diminished IL-1β secretion, respectively. Furthermore, DP1 signaling in human macrophages also resulted in the up-regulation of a putative functional analog, POP3, suggesting that PGD2 similarly modulates inflammasomes in human cells. These findings demonstrate a previously undescribed role for prostaglandin signaling in preventing excessive inflammasome activation and together with previously published results, suggest that eicosanoids and inflammasomes are reciprocally regulated. [ABSTRACT FROM AUTHOR]

Despite decades of research, malaria remains a global health crisis. Current subunit vaccine approaches do not provide efficient long-term, sterilizing immunity against Plasmodium infections in humans. Conversely, whole parasite vaccinations with their larger array of target Ags have conferred long-lasting sterilizing protection to humans. Similar studies in rodent models of malaria reveal that CD8+ T cells play a critical role in liver-stage immunity after whole parasite vaccination. However, it is unknown whether all CD8+ T cell specificities elicited by whole parasite vaccination contribute to protection, an issue of great relevance for enhanced subunit vaccination. In this article, we show that robust CD8+ T cell responses of similar phenotype are mounted after prime-boost immunization against Plasmodium berghei glideosome-associated protein 5041-48- sporozoite-specific protein 20318-325-, thrombospondin-related adhesion protein (TRAP) 130-138-, or circumsporozoite protein (CSP) 252-260-derived epitopes in mice, but only CSP252-260- and TRAP130-138-specific CD8+ T cells provide sterilizing immunity and reduce liver parasite burden after sporozoite challenge. Further, CD8+ T cells specific to sporozoite surface-expressed CSP and TRAP proteins, but not intracellular glideosome-associated protein 50 and sporozoite-specific protein 20, efficiently recognize sporozoite-infected hepatocytes in vitro. These results suggest that: 1) protection-relevant antigenic targets, regardless of their immunogenic potential, must be efficiently presented by infected hepatocytes for CD8+ T cells to eliminate liver-stage Plasmodium infection; and 2) proteins expressed on the surface of sporozoites may be good target Ags for protective CD8+ T cells. [ABSTRACT FROM AUTHOR]

Plasmodium parasites undergo development and replication within the hepatocytes before infecting the erythrocytes and initiating clinical malaria. Although type-I interferons (IFNs) are known to hinder Plasmodium infection within the liver, the underlying mechanisms remain unclear. Here, we describe two IFN-I-driven hepatocyte antimicrobial programs controlling liver-stage malaria. First, oxidative defense by NADPH oxidases 2 and 4 triggers a pathway of lysosomal fusion with the parasitophorous vacuole (PV) to help clear Plasmodium . Second, guanylate-binding protein (GBP) 1 disruption of the PV activates caspase-1 inflammasome, inducing pyroptosis to remove the infected host cells. Remarkably, both human and mouse hepatocytes enlist these cell-autonomous immune programs to eliminate Plasmodium ; their pharmacologic or genetic inhibition led to profound malarial susceptibility, and are essential in vivo . In addition to identifying the IFN-I-mediated cell-autonomous immune circuits controlling Plasmodium infection in the hepatocytes, this study extends our understanding of how non-immune cells are integral to protective immunity against malaria.

Inflammasome-mediated caspase-1 activation facilitates innate immune control of Plasmodium in the liver, thereby limiting the incidence and severity of clinical malaria. However, caspase-1 processing occurs incompletely in both mouse and human hepatocytes and precludes the generation of mature IL-1β or IL-18, unlike in other cells. Why this is so or how it impacts Plasmodium control in the liver has remained unknown. We show that an inherently reduced expression of the inflammasome adaptor molecule apoptosis-associated specklike protein containing CARD (ASC) is responsible for the incomplete proteolytic processing of caspase-1 in murine hepatocytes. Transgenically enhancing ASC expression in hepatocytes enabled complete caspase-1 processing, enhanced pyroptotic cell death, maturation of the proinflammatory cytokines IL-1β and IL-18 that was otherwise absent, and better overall control of Plasmodium infection in the liver of mice. This, however, impeded the protection offered by live attenuated antimalarial vaccination. Tempering ASC expression in mouse macrophages, on the other hand, resulted in incomplete processing of caspase-1. Our work shows how caspase-1 activation and function in host cells are fundamentally defined by ASC expression and offers a potential new pathway to create better disease and vaccination outcomes by modifying the latter., (Copyright © 2024 by The American Association of Immunologists, Inc.)

Activated CD8 + T cells differentiate into cytotoxic effector (T EFF ) cells that eliminate target cells. How T EFF cell identity is established and maintained is not fully understood. We found that Runx3 deficiency limited clonal expansion and impaired upregulation of cytotoxic molecules in T EFF cells. Runx3-deficient CD8 + T EFF cells aberrantly upregulated genes characteristic of follicular helper T (T FH ) cell lineage, including Bcl6, Tcf7 and Cxcr5. Mechanistically, the Runx3-CBFβ transcription factor complex deployed H3K27me3 to Bcl6 and Tcf7 genes to suppress the T FH program. Ablating Tcf7 in Runx3-deficient CD8 + T EFF cells prevented the upregulation of T FH genes and ameliorated their defective induction of cytotoxic genes. As such, Runx3-mediated Tcf7 repression coordinately enforced acquisition of cytotoxic functions and protected the cytotoxic lineage integrity by preventing T FH -lineage deviation.

Unlabelled: Trypanosoma cruzi is a protozoan parasite of humans and animals, affecting 10 to 20 million people and innumerable animals, primarily in the Americas. Despite being the largest cause of infection-induced heart disease worldwide, even among the neglected tropical diseases (NTDs) T. cruzi is considered one of the least well understood and understudied. The genetic complexity of T. cruzi as well as the limited set of efficient techniques for genome engineering contribute significantly to the relative lack of progress in and understanding of this pathogen. Here, we adapted the CRISPR-Cas9 system for the genetic engineering of T. cruzi, demonstrating rapid and efficient knockout of multiple endogenous genes, including essential genes. We observed that in the absence of a template, repair of the Cas9-induced double-stranded breaks (DSBs) in T. cruzi occurs exclusively by microhomology-mediated end joining (MMEJ) with various-sized deletions. When a template for DNA repair is provided, DSB repair by homologous recombination is achieved at an efficiency several orders of magnitude higher than that in the absence of CRISPR-Cas9-induced DSBs. We also demonstrate the high multiplexing capacity of CRISPR-Cas9 in T. cruzi by knocking down expression of an enzyme gene family consisting of 65 members, resulting in a significant reduction of enzymatic product with no apparent off-target mutations. Lastly, we show that Cas9 can mediate disruption of its own coding sequence, rescuing a growth defect in stable Cas9-expressing parasites. These results establish a powerful new tool for the analysis of gene functions in T. cruzi, enabling the study of essential genes and their functions and analysis of the many large families of related genes that occupy a substantial portion of the T. cruzi genome., Importance: Trypanosoma cruzi, the causative agent of human Chagas disease, is the leading worldwide cause of infectious myocarditis. Diagnostics for the infection are relatively poor, treatment options are limited and of variable effectiveness, and suitable vaccines are nonexistent. The T. cruzi genome is replete with genes of unknown function and greatly expanded gene families with hundreds of members. The absence of facile genetic engineering tools, including RNA interference, for T. cruzi has prevented elucidation of gene and gene family function and the development of better infection prevention and control measures. In this study, we demonstrate that the CRISPR-Cas9 system is a versatile and powerful tool for genome manipulations in T. cruzi, bringing new opportunities for unraveling the functions of previously uncharacterized genes and how this human pathogen engages its large families of genes encoding surface proteins to interact with human and animal hosts., (Copyright © 2014 Peng et al.)