FAM21 is critical for TLR2/CLEC4E-mediated dendritic cell function against Candida albicans.

FAM21 (family with sequence similarity 21) is a component of the Wiskott-Aldrich syndrome protein and SCAR homologue (WASH) protein complex that mediates actin polymerization at endosomal membranes to facilitate sorting of cargo-containing vesicles out of endosomes. To study the function of FAM21 in vivo, we generated conditional knockout (cKO) mice in the C57BL/6 background in which FAM21 was specifically knocked out of CD11c-positive dendritic cells. BMDCs from those mice displayed enlarged early endosomes, and altered cell migration and morphology relative to WT cells. FAM21-cKO cells were less competent in phagocytosis and protein antigen presentation in vitro, though peptide antigen presentation was not affected. More importantly, we identified the TLR2/CLEC4E signaling pathway as being down-regulated in FAM21-cKO BMDCs when challenged with its specific ligand Candida albicans Moreover, FAM21-cKO mice were more susceptible to C. albicans infection than WT mice. Reconstitution of WT BMDCs in FAM21-cKO mice rescued them from lethal C. albicans infection. Thus, our study highlights the importance of FAM21 in a host immune response against a significant pathogen.

Interactions of Candida tropicalis pH-related antigen 1 with complement proteins C3, C3b, factor-H, C4BP and complement evasion.

Candida, as a part of the human microbiota, can cause opportunistic infections that are either localised or systemic candidiasis. Emerging resistance to the standard antifungal drugs is associated with increased mortality rate due to invasive Candida infections, particularly in immunocompromised patients. While there are several species of Candida, an increasing number of Candida tropicalis isolates have been recently reported from patients with invasive candidiasis or inflammatory bowel diseases. In order to establish infections, C. tropicalis has to adopt several strategies to escape the host immune attack. Understanding the immune evasion strategies is of great importance as these can be exploited as novel therapeutic targets. C. albicans pH-related antigen 1 (CaPra1), a surface bound and secretory protein, has been found to interact strongly with the immune system and help in complement evasion. However, the role of C. tropicalis Pra1 (CtPra1) and its interaction with the complement is not studied yet. Thus, we characterised how pH-related antigen 1 of C. tropicalis (CtPra1) interacts with some of the key complement proteins of the innate immune system. CtPra1 was recombinantly produced using a Kluyveromyces lactis yeast expression system. Recombinant CtPra1, was found to bind human C3 and C3b, central molecules of the complement pathways that are important components of the innate immune system. It was also found to bind human complement regulatory proteins factor-H and C4b-binding protein (C4BP). CtPra1-factor-H and CtPra1-C4BP interactions were found to be ionic in nature as the binding intensity affected by high sodium chloride concentrations. CtPra1 inhibited functional complement activation with different effects on classical (∼20 %), lectin (∼25 %) and alternative (∼30 %) pathways. qPCR experiments using C. tropicalis clinical isolates (oral, blood and peritoneal fluid) revealed relatively higher levels of expression of CtPra1 gene when compared to the reference strain. Native CtPra1 was found to be expressed both as membrane-bound and secretory forms in the clinical isolates. Thus, C. tropicalis appears to be a master of immune evasion by using Pra1 protein. Further investigation using in-vivo models will help ascertain if these proteins can be novel therapeutic targets.

Measurements of Treg Cell Induction by Candida albicans DNA Using Flow Cytometry.

Microbiota and their metabolites in the human gut regulate a variety of immune cells including regulatory T cells (Treg cells) and cytokine production. The T helper 17 (Th17)/Treg ratio biomarker (Th17/Treg), for example, has been linked to the development and progression of certain inflammatory diseases, insulin resistance, and systemic lupus erythematosus (SLE). Candida albicans is an opportunistic fungal pathogen that also colonizes the gut. T cell reactivity through T helper cells play critical roles in fungal clearance by the host through the secretion of proinflammatory cytokines. While these cytokines are mainly produced by the Th1 and Th17 subsets of T cells, another subset of T cells, the Treg cells, are also induced by antigenic ligands from pathogens that inhibit the responses of other effector T cells during the inflammation. The antigenic ligands for Treg induction have been found to include microbial cell wall polysaccharides (PSA), metabolites like short chain fatty acids (SCFA), or even microbial DNA.

Fungal sensing enhances neutrophil metabolic fitness by regulating antifungal Glut1 activity.

Combating fungal pathogens poses metabolic challenges for neutrophils, key innate cells in anti-Candida albicans immunity, yet how host-pathogen interactions cause remodeling of the neutrophil metabolism is unclear. We show that neutrophils mediate renal immunity to disseminated candidiasis by upregulating glucose uptake via selective expression of glucose transporter 1 (Glut1). Mechanistically, dectin-1-mediated recognition of ß-glucan leads to activation of PKCδ, which triggers phosphorylation, localization, and early glucose transport by a pool of pre-formed Glut1 in neutrophils. These events are followed by increased Glut1 gene transcription, leading to more sustained Glut1 accumulation, which is also dependent on the ß-glucan/dectin-1/CARD9 axis. Card9-deficient neutrophils show diminished glucose incorporation in candidiasis. Neutrophil-specific Glut1-ablated mice exhibit increased mortality in candidiasis caused by compromised neutrophil phagocytosis, reactive oxygen species (ROS), and neutrophil extracellular trap (NET) formation. In human neutrophils, ß-glucan triggers metabolic remodeling and enhances candidacidal function. Our data show that the host-pathogen interface increases glycolytic activity in neutrophils by regulating Glut1 expression, localization, and function.

TLR Signaling Rescues Fungicidal Activity in Syk-Deficient Neutrophils.

An impaired neutrophil response to pathogenic fungi puts patients at risk for fungal infections with a high risk of morbidity and mortality. Acquired neutrophil dysfunction in the setting of iatrogenic immune modulators can include the inhibition of critical kinases such as spleen tyrosine kinase (Syk). In this study, we used an established system of conditionally immortalized mouse neutrophil progenitors to investigate the ability to augment Syk-deficient neutrophil function against Candida albicans with TLR agonist signaling. LPS, a known immunomodulatory molecule derived from Gram-negative bacteria, was capable of rescuing effector functions of Syk-deficient neutrophils, which are known to have poor fungicidal activity against Candida species. LPS priming of Syk-deficient mouse neutrophils demonstrates partial rescue of fungicidal activity, including phagocytosis, degranulation, and neutrophil swarming, but not reactive oxygen species production against C. albicans, in part due to c-Fos activation. Similarly, LPS priming of human neutrophils rescues fungicidal activity in the presence of pharmacologic inhibition of Syk and Bruton's tyrosine kinase (Btk), both critical kinases in the innate immune response to fungi. In vivo, neutropenic mice were reconstituted with wild-type or Syk-deficient neutrophils and challenged i.p. with C. albicans. In this model, LPS improved wild-type neutrophil homing to the fungal challenge, although Syk-deficient neutrophils did not persist in vivo, speaking to its crucial role on in vivo persistence. Taken together, we identify TLR signaling as an alternate activation pathway capable of partially restoring neutrophil effector function against Candida in a Syk-independent manner.

IL-33 Coordinates Innate Defense to Systemic Candida albicans Infection by Regulating IL-23 and IL-10 in an Opposite Way.

Invasive candidiasis has high mortality rates in immunocompromised patients, causing serious health problems. In mouse models, innate immunity protects the host by rapidly mobilizing a variety of resistance and tolerance mechanisms to systemic Candida albicans infection. We have previously demonstrated that exogenous IL-33 regulates multiple steps of innate immunity involving resistance and tolerance processes. In this study, we systematically analyzed the in vivo functions of endogenous IL-33 using Il33 -/- mice and in vitro immune cell culture. Tubular epithelial cells mainly secreted IL-33 in response to systemic C. albicans infection. Il33 -/- mice showed increased mortality and morbidity, which were due to impaired fungal clearance. IL-33 initiated an innate defense mechanism by costimulating dendritic cells to produce IL-23 after systemic C. albicans infection, which in turn promoted the phagocytosis of neutrophils through secretion of GM-CSF by NK cells. The susceptibility of Il33 -/- mice was also associated with increased levels of IL-10, and neutralization of IL-10 resulted in enhanced fungal clearance in Il33 -/- mice. However, depletion of IL-10 overrode the effect of IL-33 on fungal clearance. In Il10 -/- mouse kidneys, MHC class II+F4/80+ macrophages were massively differentiated after C. albicans infection, and these cells were superior to MHC class II-F4/80+ macrophages that were preferentially differentiated in wild-type mouse kidneys in killing of extracellular hyphal C. albicans Taken together, our results identify IL-33 as critical early regulator controlling a serial downstream signaling events of innate defense to C. albicans infection.

Control of ß-glucan exposure by the endo-1,3-glucanase Eng1 in Candida albicans modulates virulence.

Candida albicans is a major opportunistic pathogen of humans. It can grow as morphologically distinct yeast, pseudohyphae and hyphae, and the ability to switch reversibly among different forms is critical for its virulence. The relationship between morphogenesis and innate immune recognition is not quite clear. Dectin-1 is a major C-type lectin receptor that recognizes ß-glucan in the fungal cell wall. C. albicans ß-glucan is usually masked by the outer mannan layer of the cell wall. Whether and how ß-glucan masking is differentially regulated during hyphal morphogenesis is not fully understood. Here we show that the endo-1,3-glucanase Eng1 is differentially expressed in yeast, and together with Yeast Wall Protein 1 (Ywp1), regulates ß-glucan exposure and Dectin-1-dependent immune activation of macrophage by yeast cells. ENG1 deletion results in enhanced Dectin-1 binding at the septa of yeast cells; while eng1 ywp1 yeast cells show strong overall Dectin-1 binding similar to hyphae of wild-type and eng1 mutants. Correlatively, hyphae of wild-type and eng1 induced similar levels of cytokines in macrophage. ENG1 expression and Eng1-mediated ß-glucan trimming are also regulated by antifungal drugs, lactate and N-acetylglucosamine. Deletion of ENG1 modulates virulence in the mouse model of hematogenously disseminated candidiasis in a Dectin-1-dependent manner. The eng1 mutant exhibited attenuated lethality in male mice, but enhanced lethality in female mice, which was associated with a stronger renal immune response and lower fungal burden. Thus, Eng1-regulated ß-glucan exposure in yeast cells modulates the balance between immune protection and immunopathogenesis during disseminated candidiasis.

DOCK2 regulates antifungal immunity by regulating RAC GTPase activity.

Fungal infections cause ~1.5 million deaths each year worldwide, and the mortality rate of disseminated candidiasis currently exceeds that of breast cancer and malaria. The major reasons for the high mortality of candidiasis are the limited number of antifungal drugs and the emergence of drug-resistant species. Therefore, a better understanding of antifungal host defense mechanisms is crucial for the development of effective preventive and therapeutic strategies. Here, we report that DOCK2 (dedicator of cytokinesis 2) promotes indispensable antifungal innate immune signaling and proinflammatory gene expression in macrophages. DOCK2-deficient macrophages exhibit decreased RAC GTPase (Rac family small GTPase) activation and ROS (reactive oxygen species) production, which in turn attenuates the killing of intracellular fungi and the activation of downstream signaling pathways. Mechanistically, after fungal stimulation, activated SYK (spleen-associated tyrosine kinase) phosphorylates DOCK2 at tyrosine 985 and 1405, which promotes the recruitment and activation of RAC GTPases and then increases ROS production and downstream signaling activation. Importantly, nanoparticle-mediated delivery of in vitro transcribed (IVT) Rac1 mRNA promotes the activity of Rac1 and helps to eliminate fungal infection in vivo. Taken together, this study not only identifies a critical role of DOCK2 in antifungal immunity via regulation of RAC GTPase activity but also provides proof of concept for the treatment of invasive fungal infections by using IVT mRNA.

Study on the mechanism of Yupingfeng powder in the treatment of immunosuppression based on UPLC⁃QTOF⁃MS, network pharmacology and molecular biology verification.

AIMS: The current study aims to investigate the effect of Yupingfeng (YPF) powder on immunosuppression, and explore the possible mechanisms. MAIN METHODS: Firstly, the monomer components of YPF powder were analyzed by UPLC-QTOF-MS combined with UNIFI automatic analysis platform, then the mechanism of YPF on immunosuppressive treatment was investigated using network pharmacological method, and finally the prediction was verified in a Candida albicans (Can)-induced immunosuppressive BALB/c mouse model. KEY FINDINGS: 98 monomer compounds in YPF were obtained. Through virtual analysis and screening on the oral utilization and drug likeness properties of the components, 47 effective components were got. 9 core targets obtained were enriched in IL-17 signaling pathway. In the mouse model, YPF could reduce the number of Can and alleviate Can-induced inflammation in the kidney effectively, upregulate Can-induced low proportion of CD4+/CD8+ of splenic lymphocytes, and increase Can-induced low activity of IL-17 pathway. SIGNIFICANCE: These results demonstrate that YPF could improve the immunity of Can-induced immunosuppression in BALB/c mice through upregulating the activity of IL-17 pathway.

Interactions of Both Pathogenic and Nonpathogenic CUG Clade Candida Species with Macrophages Share a Conserved Transcriptional Landscape.

Candida species are a leading cause of opportunistic, hospital-associated bloodstream infections with high mortality rates, typically in immunocompromised patients. Several species, including Candida albicans, the most prevalent cause of infection, belong to the monophyletic CUG clade of yeasts. Innate immune cells such as macrophages are crucial for controlling infection, and C. albicans responds to phagocytosis by a coordinated induction of pathways involved in catabolism of nonglucose carbon sources, termed alternative carbon metabolism, which together are essential for virulence. However, the interactions of other CUG clade species with macrophages have not been characterized. Here, we analyzed transcriptional responses to macrophage phagocytosis by six Candida species across a range of virulence and clinical importance. We define a core induced response common to pathogenic and nonpathogenic species alike, heavily weighted to alternative carbon metabolism. One prominent pathogen, Candida parapsilosis, showed species-specific expansion of phagocytosis-responsive genes, particularly metabolite transporters. C. albicans and Candida tropicalis, the other prominent pathogens, also had species-specific responses, but these were largely comprised of functionally uncharacterized genes. Transcriptional analysis of macrophages also demonstrated highly correlated proinflammatory transcriptional responses to different Candida species that were largely independent of fungal viability, suggesting that this response is driven by recognition of conserved cell wall components. This study significantly broadens our understanding of host interactions in CUG clade species, demonstrating that although metabolic plasticity is crucial for virulence in Candida, it alone is not sufficient to confer pathogenicity. Instead, we identify sets of mostly uncharacterized genes that may explain the evolution of pathogenicity. IMPORTANCE Candidiasis is a major fungal infection by Candida species, causing life-threatening invasive disease in immunocompromised patients. C. albicans, which is adapted to commensalism of human mucosae, is the most common cause. While several other species cause infection, most are less prevalent or less virulent. As innate immune cells are the primary defense against Candida infection, we compared the transcriptional responses of C. albicans and related species to phagocytosis by macrophages, to understand the basis of variation in pathogenesis. This response, including the metabolic remodeling required for virulence in C. albicans, was strikingly conserved across the virulence spectrum. Macrophage responses to different species were also highly similar. This study indicates that important elements of host-pathogen interactions in C. albicans are not driven by adaptation to the mammalian host and improves our understanding of pathogenicity in opportunistic fungal species that are understudied but collectively impose a significant threat of their own.

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

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-1ß 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.

B Cell Recognition of Candida albicans Hyphae via TLR 2 Promotes IgG1 and IL-6 Secretion for TH17 Differentiation.

Candida albicans is usually a benign member of the human gut microbiota, but can become pathogenic under certain circumstances, for example in an immunocompromised host. The innate immune system, in particular neutrophils and macrophages, constitutes a crucial first line of defense against fungal invasion, however adaptive immunity may provide long term protection and thus allow vaccination of at risk patients. While TH1 and TH17 cells are important for antifungal responses, the role of B cells and antibodies in protection from C. albicans infection is less well defined. In this study, we show that C. albicans hyphae but not yeast, as well as fungal cell wall components, directly activate B cells via MyD88 signaling triggered by Toll- like receptor 2, leading to increased IgG1 production. While Dectin-1 signals and specific recognition by the B cell receptor are dispensable for B cell activation in this system, TLR2/MyD88 signals cooperate with CD40 signals in promoting B cell activation. Importantly, recognition of C. albicans via MyD88 signaling is also essential for induction of IL-6 secretion by B cells, which promotes TH17 polarization in T-B cell coculture experiments. B cells may thus be activated directly by C. albicans in its invasive form, leading to production of antibodies and T cell help for fungal clearance.

The network interplay of interferon and Toll-like receptor signaling pathways in the anti-Candida immune response.

Fungal infections represent a major global health problem affecting over a billion people that kills more than 1.5 million annually. In this study, we employed an integrative approach to reveal the landscape of the human immune responses to Candida spp. through meta-analysis of microarray, bulk, and single-cell RNA sequencing (scRNA-seq) data for the blood transcriptome. We identified across these different studies a consistent interconnected network interplay of signaling molecules involved in both Toll-like receptor (TLR) and interferon (IFN) signaling cascades that is activated in response to different Candida species (C. albicans, C. auris, C. glabrata, C. parapsilosis, and C. tropicalis). Among these molecules are several types I IFN, indicating an overlap with antiviral immune responses. scRNA-seq data confirmed that genes commonly identified by the three transcriptomic methods show cell type-specific expression patterns in various innate and adaptive immune cells. These findings shed new light on the anti-Candida immune response, providing putative molecular pathways for therapeutic intervention.

Therapeutic Effect of an Antibody-Derived Peptide in a Galleria mellonella Model of Systemic Candidiasis.

The synthetic peptide T11F (TCRVDHRGLTF), with sequence identical to a fragment of the constant region of human IgM, and most of its alanine-substituted derivatives proved to possess a significant candidacidal activity in vitro. In this study, the therapeutic efficacy of T11F, D5A, the derivative most active in vitro, and F11A, characterized by a different conformation, was investigated in Galleria mellonella larvae infected with Candida albicans. A single injection of F11A and D5A derivatives, in contrast with T11F, led to a significant increase in survival of larvae injected with a lethal inoculum of C. albicans cells, in comparison with infected animals treated with saline. Peptide modulation of host immunity upon C. albicans infection was determined by hemocyte analysis and larval histology, highlighting a different immune stimulation by the studied peptides. F11A, particularly, was the most active in eliciting nodule formation, melanization and fat body activation, leading to a better control of yeast infection. Overall, the obtained data suggest a double role for F11A, able to simultaneously target the fungus and the host immune system, resulting in a more efficient pathogen clearance.

Trained Innate Immunity Induced by Vaccination with Low-Virulence Candida Species Mediates Protection against Several Forms of Fungal Sepsis via Ly6G+ Gr-1+ Leukocytes.

We recently discovered a novel form of trained innate immunity (TII) induced by low-virulence Candida species (i.e., Candida dubliniensis) that protects against lethal fungal/bacterial infection. Mice vaccinated by intraperitoneal (i.p.) inoculation are protected against lethal sepsis following Candida albicans/Staphylococcus aureus (Ca/Sa) intra-abdominal infection (IAI) or Ca bloodstream infection (BSI). The protection against IAI is mediated by long-lived Gr-1+ leukocytes as putative myeloid-derived suppressor cells (MDSCs) and not by prototypical trained macrophages. This study aimed to determine if a similar TII mechanism (Gr-1+ cell-mediated suppression of sepsis) is protective against BSI and whether this TII can also be induced following intravenous (i.v.) vaccination. For this, mice were vaccinated with low-virulence Candida strains (i.p. or i.v.), followed by lethal challenge (Ca/Sa i.p. or Ca i.v.) 14 days later, and observed for sepsis (hypothermia, sepsis scoring, and serum cytokines), organ fungal burden, and mortality. Similar parameters were monitored following depletion of macrophages or Gr-1+ leukocytes during lethal challenge. The results showed that mice vaccinated i.p. or i.v. were protected against lethal Ca/Sa IAI or Ca BSI. In all cases, protection was mediated by Ly6G+ Gr-1+ putative granulocytic MDSCs (G-MDSCs), with no role for macrophages, and correlated with reduced sepsis parameters. Protection also correlated with reduced fungal burden in spleen and brain but not liver or kidney. These results suggest that Ly6G+ G-MDSC-mediated TII is induced by either the i.p. and i.v. route of inoculation and protects against IAI or BSI forms of systemic candidiasis, with survival correlating with amelioration of sepsis and reduced organ-specific fungal burden. IMPORTANCE Trained innate immunity (TII) is induced following immunization with live attenuated microbes and represents a clinically important strategy to enhance innate defenses. TII was initially demonstrated following intravenous inoculation with low-virulence Candida albicans, with protection against a subsequent lethal C. albicans intravenous bloodstream infection (BSI) mediated by monocytes with enhanced cytokine responses. We expanded this by describing a novel form of TII induced by intraperitoneal inoculation with low-virulence Candida that protects against lethal sepsis induced by polymicrobial intra-abdominal infection (IAI) via Gr-1+ leukocytes as putative myeloid-derived suppressor cells (MDSCs). In this study, we addressed these two scenarios and confirmed an exclusive role for Ly6G+ Gr-1+ leukocytes in mediating TII against either IAI or BSI via either route of inoculation, with protection associated with suppression of sepsis. These studies highlight the previously unrecognized importance of Ly6G+ MDSCs as central mediators of a novel form of TII termed trained tolerogenic immunity.

Transmission of trained immunity and heterologous resistance to infections across generations.

Intergenerational inheritance of immune traits linked to epigenetic modifications has been demonstrated in plants and invertebrates. Here we provide evidence for transmission of trained immunity across generations to murine progeny that survived a sublethal systemic infection with Candida albicans or a zymosan challenge. The progeny of trained mice exhibited cellular, developmental, transcriptional and epigenetic changes associated with the bone marrow-resident myeloid effector and progenitor cell compartment. Moreover, the progeny of trained mice showed enhanced responsiveness to endotoxin challenge, alongside improved protection against systemic heterologous Escherichia coli and Listeria monocytogenes infections. Sperm DNA of parental male mice intravenously infected with the fungus C. albicans showed DNA methylation differences linked to immune gene loci. These results provide evidence for inheritance of trained immunity in mammals, enhancing protection against infections.

Candida albicans elicits protective allergic responses via platelet mediated T helper 2 and T helper 17 cell polarization.

Fungal airway infection (airway mycosis) is an important cause of allergic airway diseases such as asthma, but the mechanisms by which fungi trigger asthmatic reactions are poorly understood. Here, we leverage wild-type and mutant Candida albicans to determine how this common fungus elicits characteristic Th2 and Th17 cell-dependent allergic airway disease in mice. We demonstrate that rather than proteinases that are essential virulence factors for molds, C. albicans instead promoted allergic airway disease through the peptide toxin candidalysin. Candidalysin activated platelets through the Von Willebrand factor (VWF) receptor GP1bα to release the Wnt antagonist Dickkopf-1 (Dkk-1) to drive Th2 and Th17 cell responses that correlated with reduced lung fungal burdens. Platelets simultaneously precluded lethal pulmonary hemorrhage resulting from fungal lung invasion. Thus, in addition to hemostasis, platelets promoted protection against C. albicans airway mycosis through an antifungal pathway involving candidalysin, GP1bα, and Dkk-1 that promotes Th2 and Th17 responses.

Signaling through Syk or CARD9 Mediates Species-Specific Anti-Candida Protection in Bone Marrow Chimeric Mice.

The spleen tyrosine kinase (Syk) and the downstream adaptor protein CARD9 are crucial signaling molecules in antimicrobial immunity. Candida parapsilosis is an emerging fungal pathogen with a high incidence in neonates, while Candida albicans is the most common agent of candidiasis. While signaling through Syk/CARD9 promotes protective host mechanisms in response to C. albicans, its function in immunity against C. parapsilosis remains unclear. Here, we generated Syk-/- and CARD9-/- bone marrow chimeric mice to study the role of Syk/CARD9 signaling in immune responses to C. parapsilosis compared to C. albicans. We demonstrate various functions of this pathway (e.g., phagocytosis, phagosome acidification, and killing) in Candida-challenged, bone marrow-derived macrophages with differential involvement of Syk and CARD9 along with species-specific differences in cytokine production. We report that Syk-/- or CARD9-/- chimeras rapidly display high susceptibility to C. albicans, while C. parapsilosis infection exacerbates over a prolonged period in these animals. Thus, our results establish that Syk and CARD9 contribute to systemic resistance to C. parapsilosis and C. albicans differently. Additionally, we confirm prior studies but also detail new insights into the fundamental roles of both proteins in immunity against C. albicans. Our data further suggest that Syk has a more prominent influence on anti-Candida immunity than CARD9. Therefore, this study reinforces the Syk/CARD9 pathway as a potential target for anti-Candida immune therapy. IMPORTANCE While C. albicans remains the most clinically significant Candida species, C. parapsilosis is an emerging pathogen with increased affinity to neonates. Syk/CARD9 signaling is crucial in immunity to C. albicans, but its role in in vivo responses to other pathogenic Candida species is largely unexplored. We used mice with hematopoietic systems deficient in Syk or CARD9 to comparatively study the function of these proteins in anti-Candida immunity. We demonstrate that Syk/CARD9 signaling has a protective role against C. parapsilosis differently than against C. albicans. Thus, this study is the first to reveal that Syk can exert immune responses during systemic Candida infections species specifically. Additionally, Syk-dependent immunity to a nonalbicans Candida species in an in vivo murine model has not been reported previously. We highlight that the contribution of Syk and CARD9 to fungal infections are not identical and underline this pathway as a promising immune-therapeutic target to fight Candida infections.

Prenatal maternal infection promotes tissue-specific immunity and inflammation in offspring.

The immune system has evolved in the face of microbial exposure. How maternal infection experienced at distinct developmental stages shapes the offspring immune system remains poorly understood. Here, we show that during pregnancy, maternally restricted infection can have permanent and tissue-specific impacts on offspring immunity. Mechanistically, maternal interleukin-6 produced in response to infection can directly impose epigenetic changes on fetal intestinal epithelial stem cells, leading to long-lasting impacts on intestinal immune homeostasis. As a result, offspring of previously infected dams develop enhanced protective immunity to gut infection and increased inflammation in the context of colitis. Thus, maternal infection can be coopted by the fetus to promote long-term, tissue-specific fitness, a phenomenon that may come at the cost of predisposition to inflammatory disorders.

Activation of Cph1 causes ß(1,3)-glucan unmasking in Candida albicans and attenuates virulence in mice in a neutrophil-dependent manner.

Masking the immunogenic cell wall epitope ß(1,3)-glucan under an outer layer of mannosylated glycoproteins is an important virulence factor deployed by Candida albicans during infection. Consequently, increased ß(1,3)-glucan exposure (unmasking) reveals C. albicans to the host's immune system and attenuates its virulence. We have previously shown that activation of the Cek1 MAPK pathway via expression of a hyperactive allele of an upstream kinase (STE11ΔN467) induced unmasking. It also increased survival of mice in a murine disseminated candidiasis model and attenuated kidney fungal burden by ≥33 fold. In this communication, we utilized cyclophosphamide-induced immunosuppression to test if the clearance of the unmasked STE11ΔN467 mutant was dependent on the host immune system. Suppression of the immune response by cyclophosphamide reduced the attenuation in fungal burden caused by the STE11ΔN467 allele. Moreover, specific depletion of neutrophils via 1A8 antibody treatment also reduced STE11ΔN467-dependent fungal burden attenuation, but to a lesser extent than cyclophosphamide, demonstrating an important role for neutrophils in mediating fungal clearance of unmasked STE11ΔN467 cells. In an effort to understand the mechanism by which Ste11ΔN467 causes unmasking, transcriptomics were used to reveal that several components in the Cek1 MAPK pathway were upregulated, including the transcription factor CPH1 and the cell wall sensor DFI1. In this report we show that a cph1ΔΔ mutation restored ß(1,3)-glucan exposure to wild-type levels in the STE11ΔN467 strain, confirming that Cph1 is the transcription factor mediating Ste11ΔN467-induced unmasking. Furthermore, Cph1 is shown to induce a positive feedback loop that increases Cek1 activation. In addition, full unmasking by STE11ΔN467 is dependent on the upstream cell wall sensor DFI1. However, while deletion of DFI1 significantly reduced Ste11ΔN467-induced unmasking, it did not impact activation of the downstream kinase Cek1. Thus, it appears that once stimulated by Ste11ΔN467, Dfi1 activates a parallel signaling pathway that is involved in Ste11ΔN467-induced unmasking.