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Regulatory T (Treg) cells are essential for the maintenance of immunological tolerance, yet the molecular components required for their maintenance and effector functions remain incompletely defined. Inactivation of VPS34 in Treg cells led to an early, lethal phenotype, with massive effector T cell activation and inflammation, like mice lacking Treg cells completely. However, VPS34-deficient Treg cells developed normally, populated the peripheral lymphoid organs and effectively supressed conventional T cells in vitro. Our data suggest that VPS34 is required for the maintaining normal numbers of mature Treg. Functionally, we observed that lack of VPS34 activity impairs cargo processing upon transendocytosis, that defective autophagy may contribute to, but is not sufficient to explain this lethal phenotype, and that loss of VPS34 activity induces a state of heightened metabolic activity that may interfere with metabolic networks required for maintenance or suppressive functions of Treg cells. [ABSTRACT FROM AUTHOR]

Background: Defective intestinal epithelial tight junction (TJ), characterized by an increase in intestinal TJ permeability, has been shown to play a critical role in the pathogenesis of inflammatory bowel disease (IBD). Tumor necrosis factor-a (TNF-α) is a key pro-inflammatory cytokine involved in the immunopathology of IBD and has been shown to cause an increase in intestinal epithelial TJ permeability. Although TNF-α antibodies and other biologics have been advanced for use in IBD treatment, these therapies are associated with severe side effects and have limited efficacy, and there is an urgent need for therapieswith benign profiles and high therapeutic efficacy. Probiotic bacteria have beneficial effects and are generally safe and represent an important class of potential therapeutic agents in IBD. Lactobacillus acidophilus (LA) is one of the most used probiotics for wide-ranging health benefits, including in gastrointestinal, metabolic, and inflammatory disorders. A specific strain of LA, LA1, was recently demonstrated to have protective and therapeutic effects on the intestinal epithelial TJ barrier. However, the mechanisms of actions of LA1 remain largely unknown. Methods: The primary aim of this study was to investigate microbial-epithelial interactions and novel signaling pathways that regulate the effect of LA1 on TNFa- induced increase in intestinal epithelial TJ permeability, using cell culture and animal model systems. Results and Conclusion: Pre-treatment of filter-grown Caco-2 monolayers with LA1 prevented the TNF-α-induced increase in intestinal epithelial TJ permeability by inhibiting TNF-α-induced activation of NF-kB p50/p65 and myosin light chain kinase (MLCK) gene and kinase activity in a TLR-2-dependent manner. LA1 produced a TLR-2- and MyD88-dependent activation of NF-kB p50/p65 in immune cells; however, LA1, in intestinal cells, inhibited the NF-kB p50/p65 activation in a TLR-2-dependent but MyD88-independent manner. In addition, LA1 inhibition of NF-kB p50/p65 and MLCK gene was mediated by TLR-2 pathway activation of phosphatidylinositol 3-kinase (PI3K) and IKK-a phosphorylation. Our results demonstrated novel intracellular signaling pathways by which LA1/TLR-2 suppresses the TNF-α pathway activation of NF-kB p50/p65 in intestinal epithelial cells and protects against the TNF-α-induced increase in intestinal epithelial TJ permeability. [ABSTRACT FROM AUTHOR]

Regulatory T (Treg) cells are essential for the maintenance of immunological tolerance, yet the molecular components required for their maintenance and effector functions remain incompletely defined. Inactivation of VPS34 in Treg cells led to an early, lethal phenotype, with massive effector T cell activation and inflammation, like mice lacking Treg cells completely. However, VPS34-deficient Treg cells developed normally, populated the peripheral lymphoid organs and effectively supressed conventional T cells in vitro. Our data suggest that VPS34 is required for the maintaining normal numbers of mature Treg. Functionally, we observed that lack of VPS34 activity impairs cargo processing upon transendocytosis, that defective autophagy may contribute to, but is not sufficient to explain this lethal phenotype, and that loss of VPS34 activity induces a state of heightened metabolic activity that may interfere with metabolic networks required for maintenance or suppressive functions of Treg cells.

BackgroundDefective intestinal epithelial tight junction (TJ), characterized by an increase in intestinal TJ permeability, has been shown to play a critical role in the pathogenesis of inflammatory bowel disease (IBD). Tumor necrosis factor-α (TNF-α) is a key pro-inflammatory cytokine involved in the immunopathology of IBD and has been shown to cause an increase in intestinal epithelial TJ permeability. Although TNF-α antibodies and other biologics have been advanced for use in IBD treatment, these therapies are associated with severe side effects and have limited efficacy, and there is an urgent need for therapies with benign profiles and high therapeutic efficacy. Probiotic bacteria have beneficial effects and are generally safe and represent an important class of potential therapeutic agents in IBD. Lactobacillus acidophilus (LA) is one of the most used probiotics for wide-ranging health benefits, including in gastrointestinal, metabolic, and inflammatory disorders. A specific strain of LA, LA1, was recently demonstrated to have protective and therapeutic effects on the intestinal epithelial TJ barrier. However, the mechanisms of actions of LA1 remain largely unknown.MethodsThe primary aim of this study was to investigate microbial-epithelial interactions and novel signaling pathways that regulate the effect of LA1 on TNF-α-induced increase in intestinal epithelial TJ permeability, using cell culture and animal model systems.Results and ConclusionPre-treatment of filter-grown Caco-2 monolayers with LA1 prevented the TNF-α-induced increase in intestinal epithelial TJ permeability by inhibiting TNF-α-induced activation of NF-κB p50/p65 and myosin light chain kinase (MLCK) gene and kinase activity in a TLR-2-dependent manner. LA1 produced a TLR-2- and MyD88-dependent activation of NF-κB p50/p65 in immune cells; however, LA1, in intestinal cells, inhibited the NF-κB p50/p65 activation in a TLR-2-dependent but MyD88-independent manner. In addition, LA1 inhibition of NF-κB p50/p65 and MLCK gene was mediated by TLR-2 pathway activation of phosphatidylinositol 3-kinase (PI3K) and IKK-α phosphorylation. Our results demonstrated novel intracellular signaling pathways by which LA1/TLR-2 suppresses the TNF-α pathway activation of NF-κB p50/p65 in intestinal epithelial cells and protects against the TNF-α-induced increase in intestinal epithelial TJ permeability.

In the past decades, advances in the use of adoptive cellular therapy to treat cancer have led to unprecedented responses in patients with relapsed/refractory or late-stage malignancies. However, cellular exhaustion and senescence limit the efficacy of FDA-approved T-cell therapies in patients with hematologic malignancies and the widespread application of this approach in treating patients with solid tumors. Investigators are addressing the current obstacles by focusing on the manufacturing process of effector T cells, including engineering approaches and ex vivo expansion strategies to regulate T-cell differentiation. Here we reviewed the current small-molecule strategies to enhance T-cell expansion, persistence, and functionality during ex vivo manufacturing. We further discussed the synergistic benefits of the dualtargeting approaches and proposed novel vasoactive intestinal peptide receptor antagonists (VIPR-ANT) peptides as emerging candidates to enhance cell-based immunotherapy. [ABSTRACT FROM AUTHOR]

NLRC3 is a member of the pattern recognition receptors nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) family, and plays a pivotal regulatory role in modulating the activation of immune cells. In macrophages, NLRC3 inhibits the activation of the NF-kB signaling pathway, the STING/TBK1 signaling pathway, and the formation of the inflammasome. In the context of T cells immune response, NLRC3 prevents the activation of T cells by regulating the function of dendritic cells and directly influencing the function of T cells. Different from other pattern recognition receptors, NLRC3 is more closely associated with regulatory activity than pathogens recognition, it influences the fates of cells, for example, prevents proliferation, promotes apoptosis and inhibits pyroptosis. These cellular functions regulated by NLRC3 are involved in the development processes of a variety of diseases, such as infectious disease, sterile inflammatory diseases, and cancer. However, its characteristics, function and regulatory mechanism in immune response and immune-related diseases have not been addressed fully. In this review, we elaborate the potential roles of NLRC3 from several different levels, include molecular mechanism, cellular functions in the immune-related diseases. [ABSTRACT FROM AUTHOR]

Regulatory T cells (Tregs) are responsible for maintaining immune homeostasis by controlling immune responses. They can be characterized by concomitant expression of FoxP3, CD25 and inhibitory receptors such as PD-1 and CTLA-4. Tregs are key players in preventing autoimmunity and are dysregulated in cancer, where they facilitate tumor immune escape. B-cell lymphoid malignancies are a group of diseases with heterogenous molecular characteristics and clinical course. Treg levels are increased in patients with B-cell lymphoid malignancies and correlate with clinical outcomes. In this review, we discuss studies investigating Treg immunobiology in B-cell lymphoid malignancies, focusing on clinical correlations, mechanisms of accumulation, phenotype, and function. Overarching trends suggest that Tregs can be induced directly by tumor cells and recruited to the tumor microenvironment where they suppress antitumor immunity to facilitate disease progression. Further, we highlight studies showing that Tregs can be modulated by novel therapeutic agents such as immune checkpoint blockade and targeted therapies. Treg disruption by novel therapeutics may beneficially restore immune competence but has been associated with occurrence of adverse events. Strategies to achieve balance between these two outcomes will be paramount in the future to improve therapeutic efficacy and safety. [ABSTRACT FROM AUTHOR]

Fibrinogen is a large molecule synthesized in the liver and released in the blood. Circulating levels of fibrinogen are upregulated after bleeding or clotting events and support wound healing. In the context of an injury, thrombin activation drives conversion of fibrinogen to fibrin. Fibrin deposition contains tissue damage, stops blood loss, and prevents microbial infection. In most circumstances, fibrin needs to be removed to allow the resolution of inflammation and tissue repair, whereas failure of this may lead to the development of various disorders. However, the contribution of fibrinogen to tissue inflammation and repair is likely to be context-dependent. In this study, the concept that fibrin needs to be removed to allow tissue repair and to reduce inflammation is challenged by our observations that, in the intestine, fibrinogen is constitutively produced by a subset of intestinal epithelial cells and deposited at the basement membrane as fibrin where it serves as a substrate for wound healing under physiological conditions such as epithelial shedding at the tip of the small intestinal villus and surface epithelium of the colon as well as under pathological conditions that require rapid epithelial repair. The functional integrity of the intestine is ensured by the constant renewal of its simple epithelium. Superficial denuding of the epithelial cell layer occurs regularly and is rapidly corrected by a process called restitution that can be influenced by various soluble and insoluble factors. Epithelial cell interaction with the extracellular matrix greatly influences the healing process by acting on cell morphology, adhesion, and migration. The functional contribution of a fibrin(ogen) matrix in the intestine was studied under physiological and pathological contexts. Our results (immunofluorescence, immunoelectron microscopy, and quantitative PCR) show that fibrin(ogen) is a novel component of the basement membrane associated with the differentiated epithelial cell population in both the small intestine and colon. Fibrin(ogen) alone is a weak ligand for epithelial cells and behaves as an anti-adhesive molecule in the presence of type I collagen. Furthermore, the presence of fibrin(ogen) significantly shortens the time required to achieve closure of wounded epithelial cell monolayers and co-cultures in a PI3K-dependent manner. In human specimens with Crohn's disease, we observed a major accumulation of fibrin(ogen) throughout the tissue and at denuded sites. In mice in which fibrin formation was inhibited with dabigatran treatment, dextran sulfate sodium administration provoked a significant increase in the disease activity index and pathological features such as mucosal ulceration and crypt abscess formation. Taken together, these results suggest that fibrin(ogen) contributes to epithelial healing under both normal and pathological conditions. [ABSTRACT FROM AUTHOR]

Aberrant innate immune responses to the gut microbiota are causally involved in the pathogenesis of inflammatory bowel diseases (IBD). The exact triggers and main signaling pathways activating innate immune cells and how they modulate adaptive immunity in IBD is still not completely understood. Here, we report that the PI3K/PTEN signaling pathway in dendritic cells enhances IL-6 production in a model of DSS-induced colitis. This results in exacerbated Th1 cell responses and increased mortality in DC-specific PTEN knockout (PTENΔDC) animals. Depletion of the gut microbiota using antibiotics as well as blocking IL-6R signaling rescued mortality in PTENΔDC mice, whereas adoptive transfer of Flt3L-derived PTEN-/- DCs into WT recipients exacerbated DSS-induced colitis and increased mortality. Taken together, we show that the PI3K signaling pathway in dendritic cells contributes to disease pathology by promoting IL-6 mediated Th1 responses. [ABSTRACT FROM AUTHOR]

Aberrant innate immune responses to the gut microbiota are causally involved in the pathogenesis of inflammatory bowel diseases (IBD). The exact triggers and main signaling pathways activating innate immune cells and how they modulate adaptive immunity in IBD is still not completely understood. Here, we report that the PI3K/PTEN signaling pathway in dendritic cells enhances IL-6 production in a model of DSS-induced colitis. This results in exacerbated Th1 cell responses and increased mortality in DC-specific PTEN knockout (PTENΔDC) animals. Depletion of the gut microbiota using antibiotics as well as blocking IL-6R signaling rescued mortality in PTENΔDC mice, whereas adoptive transfer of Flt3L-derived PTEN-/- DCs into WT recipients exacerbated DSS-induced colitis and increased mortality. Taken together, we show that the PI3K signaling pathway in dendritic cells contributes to disease pathology by promoting IL-6 mediated Th1 responses.

Exosomes are vesicles secreted by various kinds of cells, and they are rich in cholesterol, sphingomyelin (SM), phosphatidylcholine, and phosphatidylserine. Although cellular sphingolipid-mediated exosome release has been reported, the involvement of other lipid components of cell membranes in the regulation of exosome release is poorly understood. Here, we show that the level of exosome release into conditioned media is significantly reduced in cultured astrocytes prepared from apolipoprotein E (ApoE) knock-out mice when compared to those prepared from wild-type (WT) mice. The reduced level of exosome release was accompanied by elevated levels of cellular cholesterol. The addition of cholesterol to WT astrocytes significantly increased the cellular cholesterol levels and reduced exosome release. PI3K/Akt phosphorylation was enhanced in ApoE-deficient and cholesterol-treated WT astrocytes. In contrast, the depletion of cholesterol in ApoE-deficient astrocytes due to treatment with β-cyclodextrin recovered the exosome release level to a level similar to that in WT astrocytes. In addition, the reduced levels of exosome release due to the addition of cholesterol recovered to the control levels after treatment with a PI3K inhibitor (LY294002). The cholesterol-dependent regulation of exosome release was also confirmed by in vivo experiments; that is, exosome levels were significantly reduced in the CSF and blood serum of WT mice that were fed a high-fat diet and had increased cholesterol levels when compared to those in WT mice that were fed a normal diet. These results suggest that exosome release is regulated by cellular cholesterol via stimulation of the PI3K/Akt signal pathway.

Exosomes are vesicles secreted by various kinds of cells, and they are rich in cholesterol, sphingomyelin (SM), phosphatidylcholine, and phosphatidylserine. Although cellular sphingolipid-mediated exosome release has been reported, the involvement of other lipid components of cell membranes in the regulation of exosome release is poorly understood. Here, we show that the level of exosome release into conditioned media is significantly reduced in cultured astrocytes prepared from apolipoprotein E (ApoE) knock-out mice when compared to those prepared from wild-type (WT) mice. The reduced level of exosome release was accompanied by elevated levels of cellular cholesterol. The addition of cholesterol to WT astrocytes significantly increased the cellular cholesterol levels and reduced exosome release. PI3K/Akt phosphorylation was enhanced in ApoE-deficient and cholesterol-treated WT astrocytes. In contrast, the depletion of cholesterol in ApoE-deficient astrocytes due to treatment with β-cyclodextrin recovered the exosome release level to a level similar to that in WT astrocytes. In addition, the reduced levels of exosome release due to the addition of cholesterol recovered to the control levels after treatment with a PI3K inhibitor (LY294002). The cholesterol-dependent regulation of exosome release was also confirmed by in vivo experiments; that is, exosome levels were significantly reduced in the CSF and blood serum of WT mice that were fed a high-fat diet and had increased cholesterol levels when compared to those in WT mice that were fed a normal diet. These results suggest that exosome release is regulated by cellular cholesterol via stimulation of the PI3K/Akt signal pathway. [ABSTRACT FROM AUTHOR]

PI3K signalling is required for activation, differentiation, and trafficking of T cells. PI3Kδ, the dominant PI3K isoform in T cells, has been extensively characterised using PI3Kδ mutant mouse models and PI3K inhibitors. Furthermore, characterisation of patients with Activated PI3K Delta Syndrome (APDS) and mouse models with hyperactive PI3Kδ have shed light on how increased PI3Kδ activity affects T cell functions. An important function of PI3Kδ is that it acts downstream of TCR stimulation to activate the major T cell integrin, LFA-1, which controls transendothelial migration of T cells as well as their interaction with antigen-presenting cells. PI3Kδ also suppresses the cell surface expression of CD62L and CCR7 which controls the migration of T cells across high endothelial venules in the lymph nodes and S1PR1 which controls lymph node egress. Therefore, PI3Kδ can control both entry and exit of T cells from lymph nodes as well as the recruitment to and retention of T cells within inflamed tissues. This review will focus on the regulation of adhesion receptors by PI3Kδ and how this contributes to T cell trafficking and localisation. These findings are relevant for our understanding of how PI3Kδ inhibitors may affect T cell redistribution and function.

PI3K signalling is required for activation, differentiation, and trafficking of T cells. PI3Kδ, the dominant PI3K isoform in T cells, has been extensively characterised using PI3Kδ mutant mouse models and PI3K inhibitors. Furthermore, characterisation of patients with Activated PI3K Delta Syndrome (APDS) and mouse models with hyperactive PI3Kδ have shed light on how increased PI3Kδ activity affects T cell functions. An important function of PI3Kδ is that it acts downstream of TCR stimulation to activate the major T cell integrin, LFA-1, which controls transendothelial migration of T cells as well as their interaction with antigen-presenting cells. PI3Kδ also suppresses the cell surface expression of CD62L and CCR7 which controls the migration of T cells across high endothelial venules in the lymph nodes and S1PR1 which controls lymph node egress. Therefore, PI3Kδ can control both entry and exit of T cells from lymph nodes as well as the recruitment to and retention of T cells within inflamed tissues. This review will focus on the regulation of adhesion receptors by PI3Kδ and how this contributes to T cell trafficking and localisation. These findings are relevant for our understanding of how PI3Kδ inhibitors may affect T cell redistribution and function. [ABSTRACT FROM AUTHOR]

Phosphoinositide 3-kinase p110 delta (PI3K p110δ) is pivotal for CD8+ T cell immune responses. The current study explores PI3K p110δ induction and repression of antigen receptor and cytokine regulated programs to inform how PI3K p110δ directs CD8+ T cell fate. The studies force a revision of the concept that PI3K p110δ controls metabolic pathways in T cells and reveal major differences in PI3K p110δ regulated transcriptional programs between naïve and effector cytotoxic T cells (CTL). These differences include differential control of the expression of cytolytic effector molecules and costimulatory receptors. Key insights from the work include that PI3K p110δ signalling pathways repress expression of the critical inhibitory receptors CTLA4 and SLAMF6 in CTL. Moreover, in both naïve and effector T cells the dominant role for PI3K p110δ is to restrain the production of the chemokines that orchestrate communication between adaptive and innate immune cells. The study provides a comprehensive resource for understanding how PI3K p110δ uses multiple processes mediated by Protein Kinase B/AKT, FOXO1 dependent and independent mechanisms and mitogen-activated protein kinases (MAPK) to direct CD8+ T cell fate.

Neutrophils, the most abundant circulating leukocytes in humans have key roles in host defense and in the inflammatory response. Agonist-activated phosphoinositide 3-kinases (PI3Ks) are important regulators of many facets of neutrophil biology. PIP3 is subject to dephosphorylation by several 5’ phosphatases, including SHIP family phosphatases, which convert the PI3K product and lipid second messenger phosphatidylinositol 3,4,5-trisphosphate (PIP3) into PI(3,4)P2, a lipid second messenger in its own right. In addition to the leukocyte restricted SHIP1, neutrophils express the ubiquitous SHIP2. This study analyzed mice and isolated neutrophils carrying a catalytically inactive SHIP2, identifying an important regulatory function in neutrophil chemotaxis and directionality in vitro and in neutrophil recruitment to sites of sterile inflammation in vivo, in the absence of major defects of any other neutrophil functions analyzed, including, phagocytosis and the formation of reactive oxygen species. Mechanistically, this is explained by a subtle effect on global 3-phosphorylated phosphoinositide species. This work identifies a non-redundant role for the hitherto overlooked SHIP2 in the regulation of neutrophils, and specifically, neutrophil chemotaxis/trafficking. It completes an emerging wider understanding of the complexity of PI3K signaling in the neutrophil, and the roles played by individual kinases and phosphatases within.

Phosphoinositide 3-kinase p110 delta (PI3K p110δ) is pivotal for CD8+ T cell immune responses. The current study explores PI3K p110δ induction and repression of antigen receptor and cytokine regulated programs to inform how PI3K p110δ directs CD8+ T cell fate. The studies force a revision of the concept that PI3K p110δ controls metabolic pathways in T cells and reveal major differences in PI3K p110δ regulated transcriptional programs between naïve and effector cytotoxic T cells (CTL). These differences include differential control of the expression of cytolytic effector molecules and costimulatory receptors. Key insights from the work include that PI3K p110δ signalling pathways repress expression of the critical inhibitory receptors CTLA4 and SLAMF6 in CTL. Moreover, in both naïve and effector T cells the dominant role for PI3K p110δ is to restrain the production of the chemokines that orchestrate communication between adaptive and innate immune cells. The study provides a comprehensive resource for understanding how PI3K p110δ uses multiple processes mediated by Protein Kinase B/AKT, FOXO1 dependent and independent mechanisms and mitogen-activated protein kinases (MAPK) to direct CD8+ T cell fate. [ABSTRACT FROM AUTHOR]

Neutrophils, the most abundant circulating leukocytes in humans have key roles in host defense and in the inflammatory response. Agonist-activated phosphoinositide 3-kinases (PI3Ks) are important regulators of many facets of neutrophil biology. PIP3 is subject to dephosphorylation by several 5' phosphatases, including SHIP family phosphatases, which convert the PI3K product and lipid second messenger phosphatidylinositol 3,4,5-trisphosphate (PIP3) into PI(3,4)P2, a lipid second messenger in its own right. In addition to the leukocyte restricted SHIP1, neutrophils express the ubiquitous SHIP2. This study analyzed mice and isolated neutrophils carrying a catalytically inactive SHIP2, identifying an important regulatory function in neutrophil chemotaxis and directionality in vitro and in neutrophil recruitment to sites of sterile inflammation in vivo , in the absence of major defects of any other neutrophil functions analyzed, including, phagocytosis and the formation of reactive oxygen species. Mechanistically, this is explained by a subtle effect on global 3-phosphorylated phosphoinositide species. This work identifies a non-redundant role for the hitherto overlooked SHIP2 in the regulation of neutrophils, and specifically, neutrophil chemotaxis/trafficking. It completes an emerging wider understanding of the complexity of PI3K signaling in the neutrophil, and the roles played by individual kinases and phosphatases within. [ABSTRACT FROM AUTHOR]

Activation of phosphoinositide 3-kinase (PI3K) is required for B cell proliferation and survival. PI3K signaling also controls key aspects of B cell differentiation. Upon engagement of the B cell receptor (BCR), PI3K activation promotes Ca(2+) mobilization and activation of NFκB-dependent transcription, events which are essential for B cell proliferation. PI3K also initiates a distinct signaling pathway involving the Akt and mTOR serine/threonine kinases. It has been generally assumed that activation of Akt and mTOR downstream of PI3K is essential for B cell function. However, Akt and mTOR have complex roles in B cell fate decisions and suppression of this pathway can enhance certain B cell responses while repressing others. In this review we will discuss genetic and pharmacological studies of Akt and mTOR function in normal B cells, and in malignancies of B cell origin.

Phosphoinositide 3-kinases (PI3Ks) play a central role in adaptive immunity by transducing signals from the T cell antigen receptor (TCR) via production of PIP3. PI3Kδ is a heterodimer composed of a p110δ catalytic subunit associated with a p85α or p85β regulatory subunit and is preferentially engaged by the TCR upon T cell activation. The molecular mechanisms leading to PI3Kδ recruitment and activation at the TCR signalosome remain unclear. In this study, we have used quantitative mass spectrometry, biochemical approaches and CRISPR-Cas9 gene editing to uncover the p110δ interactome in primary CD4+ T cells. Moreover, we have determined how the PI3Kδ interactome changes upon the differentiation of small naïve T cells into T cell blasts expanded in the presence of IL-2. Our interactomic analyses identified multiple constitutive and inducible PI3Kδ-interacting proteins, some of which were common to naïve and previously-activated T cells. Our data reveals that PI3Kδ rapidly interacts with as many as seven adaptor proteins upon TCR engagement, including the Gab-family proteins, GAB2 and GAB3, a CD5-CBL signalosome and the transmembrane proteins ICOS and TRIM. Our results also suggest that PI3Kδ pre-forms complexes with the adaptors SH3KBP1 and CRKL in resting cells that could facilitate the localization and activation of p110δ at the plasma membrane by forming ternary complexes during early TCR signalling. Furthermore, we identify interactions that were not previously known to occur in CD4+ T cells, involving BCAP, GAB3, IQGAP3 and JAML. We used CRISPR-Cas9-mediated gene knockout in primary T cells to confirm that BCAP is a positive regulator of PI3K-AKT signalling in CD4+ T cell blasts. Overall, our results provide evidence for a large protein network that regulates the recruitment and activation of PI3Kδ in T cells. Finally, this work shows how the PI3Kδ interactome is remodeled as CD4+ T cells differentiate from naïve T cells to activated T cell blasts. These activated T cells upregulate additional PI3Kδ adaptor proteins, including BCAP, GAB2, IQGAP3 and ICOS. This rewiring of TCR-PI3K signalling that occurs upon T cell differentiation may serve to reduce the threshold of activation and diversify the inputs for the PI3K pathway in effector T cells.

In the autoimmune disease Systemic Lupus Erythematosus (SLE), autoantibodies are formed that promote inflammation and tissue damage. There has been significant interest in understanding the B cell derangements involved in SLE pathogenesis. The past few years have been particularly fruitful in three domains: the role of PI3K signaling in loss of B cell tolerance, the role of IFNγ signaling in the development of autoimmunity, and the characterization of changes in chromatin accessibility in SLE B cells. The PI3K pathway coordinates various downstream signaling molecules involved in B cell development and activation. It is governed by the phosphatases PTEN and SHIP-1. Murine models lacking either of these phosphatases in B cells develop autoimmune disease and exhibit defects in B cell tolerance. Limited studies of human SLE B cells demonstrate reduced expression of PTEN or increased signaling events downstream of PI3K in some patients. IFNγ has long been known to be elevated in both SLE patients and mouse models of lupus. New data suggests that IFNγR expression on B cells is required to develop autoreactive germinal centers (GC) and autoantibodies in murine lupus. Furthermore, IFNγ promotes increased transcription of BCL6, IL-6 and T-bet in B cells, which also promote GC and autoantibody formation. IFNγ also induces epigenetic changes in human B cells. SLE B cells demonstrate significant epigenetic reprogramming, including enhanced chromatin accessibility at transcription factor motifs involved in B cell activation and plasma cell (PC) differentiation as well as alterations in DNA methylation and histone modifications. Histone deacetylase inhibitors limit disease development in murine lupus models, at least in part via their ability to prevent B cell class switching and differentiation into plasma cells. This review will discuss relevant discoveries of the past several years pertaining to these areas of SLE B cell biology.

The spread of the novel human respiratory coronavirus (SARS-CoV-2) is a global public health emergency. There is no known successful treatment as of this time, and there is a need for medical options to mitigate this current epidemic. SARS-CoV-2 uses the angiotensin-converting enzyme 2 (ACE2) receptor and is primarily trophic for the lower and upper respiratory tract. A number of current studies on COVID-19 have demonstrated the substantial increase in pro-inflammatory factors in the lungs during infection. The virus is also documented in the central nervous system and, particularly in the brainstem, which plays a key role in respiratory and cardiovascular function. Currently, there are few antiviral approaches, and several alternative drugs are under investigation. Two of these are Idelalisib and Ebastine, already proposed as preventive strategies in airways and allergic diseases. The interesting and evolving potential of phosphoinositide 3-kinase δ (PI3Kδ) inhibitors, together with Ebastine, lies in their ability to suppress the release of pro-inflammatory cytokines, such as IL-1β, IL-8, IL-6, and TNF-α, by T cells. This may represent an optional therapeutic choice for COVID-19 to reduce inflammatory reactions and mortality, enabling patients to recover faster. This concise communication aims to provide new potential therapeutic targets capable of mitigating and alleviating SARS-CoV-2 pandemic infection.

Phosphoinositide 3-kinases (PI3Ks) play a central role in adaptive immunity by transducing signals from the T cell antigen receptor (TCR) via production of PIP3. PI3Kδ is a heterodimer composed of a p110δ catalytic subunit associated with a p85α or p85β regulatory subunit and is preferentially engaged by the TCR upon T cell activation. The molecular mechanisms leading to PI3Kδ recruitment and activation at the TCR signalosome remain unclear. In this study, we have used quantitative mass spectrometry, biochemical approaches and CRISPR-Cas9 gene editing to uncover the p110δ interactome in primary CD4+ T cells. Moreover, we have determined how the PI3Kδ interactome changes upon the differentiation of small naïve T cells into T cell blasts expanded in the presence of IL-2. Our interactomic analyses identified multiple constitutive and inducible PI3Kδ-interacting proteins, some of which were common to naïve and previously-activated T cells. Our data reveals that PI3Kδ rapidly interacts with as many as seven adaptor proteins upon TCR engagement, including the Gab-family proteins, GAB2 and GAB3, a CD5-CBL signalosome and the transmembrane proteins ICOS and TRIM. Our results also suggest that PI3Kδ pre-forms complexes with the adaptors SH3KBP1 and CRKL in resting cells that could facilitate the localization and activation of p110δ at the plasma membrane by forming ternary complexes during early TCR signalling. Furthermore, we identify interactions that were not previously known to occur in CD4+ T cells, involving BCAP, GAB3, IQGAP3 and JAML. We used CRISPR-Cas9-mediated gene knockout in primary T cells to confirm that BCAP is a positive regulator of PI3K-AKT signalling in CD4+ T cell blasts. Overall, our results provide evidence for a large protein network that regulates the recruitment and activation of PI3Kδ in T cells. Finally, this work shows how the PI3Kδ interactome is remodeled as CD4+ T cells differentiate from naïve T cells to activated T cell blasts. These activated T cells upregulate additional PI3Kδ adaptor proteins, including BCAP, GAB2, IQGAP3 and ICOS. This rewiring of TCR-PI3K signalling that occurs upon T cell differentiation may serve to reduce the threshold of activation and diversify the inputs for the PI3K pathway in effector T cells. [ABSTRACT FROM AUTHOR]

In the autoimmune disease Systemic Lupus Erythematosus (SLE), autoantibodies are formed that promote inflammation and tissue damage. There has been significant interest in understanding the B cell derangements involved in SLE pathogenesis. The past few years have been particularly fruitful in three domains: the role of PI3K signaling in loss of B cell tolerance, the role of IFNγ signaling in the development of autoimmunity, and the characterization of changes in chromatin accessibility in SLE B cells. The PI3K pathway coordinates various downstream signaling molecules involved in B cell development and activation. It is governed by the phosphatases PTEN and SHIP-1. Murine models lacking either of these phosphatases in B cells develop autoimmune disease and exhibit defects in B cell tolerance. Limited studies of human SLE B cells demonstrate reduced expression of PTEN or increased signaling events downstream of PI3K in some patients. IFNγ has long been known to be elevated in both SLE patients and mouse models of lupus. New data suggests that IFNγR expression on B cells is required to develop autoreactive germinal centers (GC) and autoantibodies in murine lupus. Furthermore, IFNγ promotes increased transcription of BCL6, IL-6 and T-bet in B cells, which also promote GC and autoantibody formation. IFNγ also induces epigenetic changes in human B cells. SLE B cells demonstrate significant epigenetic reprogramming, including enhanced chromatin accessibility at transcription factor motifs involved in B cell activation and plasma cell (PC) differentiation as well as alterations in DNA methylation and histone modifications. Histone deacetylase inhibitors limit disease development in murine lupus models, at least in part via their ability to prevent B cell class switching and differentiation into plasma cells. This review will discuss relevant discoveries of the past several years pertaining to these areas of SLE B cell biology. [ABSTRACT FROM AUTHOR]

The spread of the novel human respiratory coronavirus (SARS-CoV-2) is a global public health emergency. There is no known successful treatment as of this time, and there is a need for medical options to mitigate this current epidemic. SARS-CoV-2 uses the angiotensin-converting enzyme 2 (ACE2) receptor and is primarily trophic for the lower and upper respiratory tract. A number of current studies on COVID-19 have demonstrated the substantial increase in pro-inflammatory factors in the lungs during infection. The virus is also documented in the central nervous system and, particularly in the brainstem, which plays a key role in respiratory and cardiovascular function. Currently, there are few antiviral approaches, and several alternative drugs are under investigation. Two of these are Idelalisib and Ebastine, already proposed as preventive strategies in airways and allergic diseases. The interesting and evolving potential of phosphoinositide 3-kinase δ (PI3Kδ) inhibitors, together with Ebastine, lies in their ability to suppress the release of pro-inflammatory cytokines, such as IL-1β, IL-8, IL-6, and TNF-α, by T cells. This may represent an optional therapeutic choice for COVID-19 to reduce inflammatory reactions and mortality, enabling patients to recover faster. This concise communication aims to provide new potential therapeutic targets capable of mitigating and alleviating SARS-CoV-2 pandemic infection. [ABSTRACT FROM AUTHOR]

SHIP-1 is an inositol phosphatase that hydrolyzes phosphatidylinositol 3-kinase (PI3K) products and negatively regulates protein kinase B (Akt) activity, thereby modulating a variety of cellular processes in mammals. However, the role of SHIP-1 in bacterial-induced sepsis is largely unknown. Here, we show that SHIP-1 regulates inflammatory responses during Gram-negative bacterium Pseudomonas aeruginosa infection. We found that infected-SHIP-1−/− mice exhibited decreased survival rates, increased inflammatory responses, and susceptibility owing to elevated expression of PI3K than wild-type (WT) mice. Inhibiting SHIP-1 via siRNA silencing resulted in lipid raft aggregates, aggravated oxidative damage, and bacterial burden in macrophages after PAO1 infection. Mechanistically, SHIP-1 deficiency augmented phosphorylation of PI3K and nuclear transcription of signal transducer and activator of transcription 5 (STAT5) to induce the expression of Trib1, which is critical for differentiation of M2 but not M1 macrophages. These findings reveal a previously unrecognized role of SHIP-1 in inflammatory responses and macrophage homeostasis during P. aeruginosa infection through a PI3K/Akt–STAT5–Trib1 axis.

Class 1 Phosphoinositide-3-Kinases (PI3Ks) have been widely studied and mediate essential roles in cellular proliferation, chemotaxis, insulin sensitivity, and immunity. Here, we provide a comprehensive overview of how macrophage expressed PI3Ks and their downstream pathways orchestrate responses to metabolic stimuli and nutrients, polarizing macrophages, shaping their cellular identity and function. Particular emphasis will be given to adipose tissue macrophages, crucial players of insulin resistance and chronic metabolically triggered inflammation during obesity. An understanding of PI3K dependent wiring of macrophage responses is important as this is involved in various diseases ranging from obesity, type 2 diabetes to chronic inflammatory disease.

Gain-of-function variants in p110δ, the catalytic subunit of phosphatidylinositol 3-kinase (PI3K) expressed in lymphocytes, cause activated PI3-kinase δ syndrome (APDS), a primary immunodeficiency that is characterized by recurrent infections, viremia, lymphadenopathy, and autoimmunity. The mechanism of autoimmunity in APDS has not been well-understood. Here, we show the profound skewing of peripheral CD4+ T cells to a T follicular helper (TFH) phenotype in a patient with APDS bearing a novel p110δ variant, Y524S. We also saw a diminishment of transient Foxp3 expression in activated T cells. Mechanistic studies revealed that both the new variant and a previously described, pathogenic variant (E81K) enhanced an interaction between intracellular Osteopontin and p85α. This interaction had been shown in mice to promote TFH differentiation. Our results demonstrate a new influence of PI3K on human T cell differentiation that is unrelated to its lipid-kinase activity and suggest that TFH should be monitored in APDS patients.

SHIP-1 is an inositol phosphatase that hydrolyzes phosphatidylinositol 3-kinase (PI3K) products and negatively regulates protein kinase B (Akt) activity, thereby modulating a variety of cellular processes in mammals. However, the role of SHIP-1 in bacterial-induced sepsis is largely unknown. Here, we show that SHIP-1 regulates inflammatory responses during Gram-negative bacterium Pseudomonas aeruginosa infection. We found that infected-SHIP-1−/− mice exhibited decreased survival rates, increased inflammatory responses, and susceptibility owing to elevated expression of PI3K than wild-type (WT) mice. Inhibiting SHIP-1 via siRNA silencing resulted in lipid raft aggregates, aggravated oxidative damage, and bacterial burden in macrophages after PAO1 infection. Mechanistically, SHIP-1 deficiency augmented phosphorylation of PI3K and nuclear transcription of signal transducer and activator of transcription 5 (STAT5) to induce the expression of Trib1, which is critical for differentiation of M2 but not M1 macrophages. These findings reveal a previously unrecognized role of SHIP-1 in inflammatory responses and macrophage homeostasis during P. aeruginosa infection through a PI3K/Akt–STAT5–Trib1 axis. [ABSTRACT FROM AUTHOR]

Phosphoinositide 3-kinase (PI3K) plays an integral role in lymphocyte function. Mutations in PIK3CD and PIK3R1, encoding the PI3K p110δ and p85α subunits, respectively, cause increased PI3K activity and result in immunodeficiency with immune dysregulation. We describe here the first cases of disseminated and congenital toxoplasmosis in a mother and child who share a pathogenic mutation in PIK3R1 and review the mechanisms underlying susceptibility to severe Toxoplasma gondii infection in activated PI3Kδ syndrome (APDS) and in other forms of primary immunodeficiency.

T follicular helper (Tfh) cells are a specialized population of CD4+ T cells that provide help to B cells for the formation and maintenance germinal centers, and the production of high affinity class-switched antibodies, long-lived plasma cells, and memory B cells. As such, Tfh cells are essential for the generation of successful long-term humoral immunity and memory responses to vaccination and infection. Conversely, overproduction of Tfh cells has been associated with the generation of autoantibodies and autoimmunity. Data from gene-targeted mice, pharmacological inhibitors, as well as studies of human and mice expressing activating mutants have revealed that PI3Kδ is a key regulator of Tfh cell differentiation, acting downstream of ICOS to facilitate inactivation of FOXO1, repression of Klf2 and induction of Bcl6. Nonetheless, here we show that after acute LCMV infection, WT and activated-PI3Kδ mice (Pik3cdE1020K/+) show comparable ratios of Tfh:Th1 viral specific CD4+ T cells, despite higher polyclonal Tfh cells in Pik3cdE1020K/+ mice. Thus, the idea that PI3K activity primarily drives Tfh cell differentiation may be an oversimplification and PI3K-mediated pathways are likely to integrate multiple signals to promote distinct effector T cell lineages. The consequences of dysregulated Tfh cell generation will be discussed in the context of the human primary immunodeficiency “Activated PI3K-delta Syndrome” (APDS), also known as “p110 delta-activating mutation causing senescent T cells, lymphadenopathy and immunodeficiency” (PASLI). Overall, these data underscore a major role for PI3K signaling in the orchestration of T lymphocyte responses.

Phosphoinositide-3 kinases (PI3Ks) generate 3-phosphorylated phosphoinositide lipids that are implicated in many biological processes in homeostatic states and pathologies such as cancer, inflammation and autoimmunity. Eight isoforms of PI3K exist in mammals and among them the class I PI3K, p110γ, and PI3Kδ, and class III Vps34 being the most expressed and well characterized in immune cells. Following engagement of pathogen recognition receptors (PRRs), PI3Ks coordinate vital cellular processes of signaling and vesicular trafficking in innate phagocytes such as macrophages and professional antigen presenting dendritic cells (DCs). Although previous studies demonstrated the involvement of PI3K isoforms in innate and adaptive immune cell types, the role of PI3Ks with respect to DC biology has been enigmatic. Thus, this review, based on studies involving PI3K isoforms, highlight how the different PI3Ks isoforms could regulate DC functions such as antigen processing and presentation including PRR responses.

Primary immunodeficiencies (PIDs) represent a group of mostly monogenic disorders caused by loss- or gain-of-function mutations in over 340 known genes that lead to abnormalities in the development and/or the function of the immune system. However, mutations in different genes can affect the same cell-signaling pathway and result in overlapping clinical phenotypes. In particular, mutations in the genes encoding for members of the phosphoinositide3-kinase (PI3K)/AKT/mTOR/S6 kinase (S6K) signaling cascade or for molecules interacting with this pathway have been associated with different PIDs that are often characterized by the coexistence of both immune deficiency and autoimmunity. The serine/threonine kinase mechanistic/mammalian target of rapamycin (mTOR), which acts downstream of PI3K and AKT, is emerging as a key regulator of immune responses. It integrates a variety of signals from the microenvironment to control cell growth, proliferation, and metabolism. mTOR plays therefore a central role in the regulation of immune cells’ differentiation and functions. Here, we review the different PIDs that share an impairment of the PI3K/AKT/mTOR/S6K pathway and we propose to name them “immune TOR-opathies” by analogy with a group of neurological disorders that has been originally defined by PB Crino and that are due to aberrant mTOR signaling (1). A better understanding of the role played by this complex intracellular cascade in the pathophysiology of “immune TOR-opathies” is crucial to develop targeted therapies.

Phosphoinositide 3-kinase (PI3K) plays an integral role in lymphocyte function. Mutations in PIK3CD and PIK3R1 , encoding the PI3K p110δ and p85α subunits, respectively, cause increased PI3K activity and result in immunodeficiency with immune dysregulation. We describe here the first cases of disseminated and congenital toxoplasmosis in a mother and child who share a pathogenic mutation in PIK3R1 and review the mechanisms underlying susceptibility to severe Toxoplasma gondii infection in activated PI3Kδ syndrome (APDS) and in other forms of primary immunodeficiency. [ABSTRACT FROM AUTHOR]

Phosphoinositide-3 kinases (PI3Ks) generate 3-phosphorylated phosphoinositide lipids that are implicated in many biological processes in homeostatic states and pathologies such as cancer, inflammation and autoimmunity. Eight isoforms of PI3K exist in mammals and among them the class I PI3K, p110γ, and PI3Kδ, and class III Vps34 being the most expressed and well characterized in immune cells. Following engagement of pathogen recognition receptors (PRRs), PI3Ks coordinate vital cellular processes of signaling and vesicular trafficking in innate phagocytes such as macrophages and professional antigen presenting dendritic cells (DCs). Although previous studies demonstrated the involvement of PI3K isoforms in innate and adaptive immune cell types, the role of PI3Ks with respect to DC biology has been enigmatic. Thus, this review, based on studies involving PI3K isoforms, highlight how the different PI3Ks isoforms could regulate DC functions such as antigen processing and presentation including PRR responses. [ABSTRACT FROM AUTHOR]

Primary immunodeficiencies (PIDs) represent a group of mostly monogenic disorders caused by loss- or gain-of-function mutations in over 340 known genes that lead to abnormalities in the development and/or the function of the immune system. However, mutations in different genes can affect the same cell-signaling pathway and result in overlapping clinical phenotypes. In particular, mutations in the genes encoding for members of the phosphoinositide3-kinase (PI3K)/AKT/mTOR/S6 kinase (S6K) signaling cascade or for molecules interacting with this pathway have been associated with different PIDs that are often characterized by the coexistence of both immune deficiency and autoimmunity. The serine/threonine kinase mechanistic/mammalian target of rapamycin (mTOR), which acts downstream of PI3K and AKT, is emerging as a key regulator of immune responses. It integrates a variety of signals from the microenvironment to control cell growth, proliferation, and metabolism. mTOR plays therefore a central role in the regulation of immune cells' differentiation and functions. Here, we review the different PIDs that share an impairment of the PI3K/AKT/mTOR/S6K pathway and we propose to name them "immune TOR-opathies" by analogy with a group of neurological disorders that has been originally defined by PB Crino and that are due to aberrant mTOR signaling (1). A better understanding of the role played by this complex intracellular cascade in the pathophysiology of "immune TOR-opathies" is crucial to develop targeted therapies. [ABSTRACT FROM AUTHOR]

Pathogenic gain-of-function mutations in the gene encoding phosphoinositide 3-kinase delta (PI3Kδ) cause activated PI3Kδ syndrome (APDS), a disease characterized by humoral immunodeficiency, lymphadenopathy, and an inability to control persistent viral infections including Epstein–Barr virus (EBV) and cytomegalovirus (CMV) infections. Understanding the mechanisms leading to impaired immune response is important to optimally treat APDS patients. Immunosenescence of CD8+ T cells was suggested to contribute to APDS pathogenesis. However, the constitutive activation of T cells in APDS may also result in T cell exhaustion. Therefore, we studied exhaustion of the CD8+ T cell compartment in APDS patients and compared them with healthy controls and HIV patients, as a control for exhaustion. The subset distribution of the T cell compartment of APDS patients was comparable with HIV patients with decreased naive CD4+ and CD8+ T cells and increased effector CD8+ T cells. Like in HIV+ patients, expression of activation markers and inhibitory receptors CD160, CD244, and programmed death receptor (PD)-1 on CD8+ T cells was increased in APDS patients, indicating exhaustion. EBV-specific CD8+ T cells from APDS patients exhibited an exhausted phenotype that resembled HIV-specific CD8+ T cells in terms of inhibitory receptor expression. Inhibition of PD-1 on EBV-specific CD8+ T cells from APDS patients enhanced in vitro proliferation and effector cytokine production. Based on these results, we conclude that total and EBV-specific CD8+ T cells from APDS patients are characterized by T cell exhaustion. Furthermore, PD-1 checkpoint inhibition may provide a possible therapeutic approach to support the immune system of APDS patients to control EBV and CMV.

Class IA phosphatidylinositol 3-kinase (PI3K) catalytic subunits p110α and p110δ are targets in cancer therapy expressed at high levels in T lymphocytes. The role of p110δ PI3K in normal or pathological immune responses is well established, yet the importance of p110α subunits in T cell-dependent immune responses is not clear. To address this problem, mice with p110α conditionally deleted in CD4+ and CD8+ T lymphocytes (p110α−/−ΔT) were used. p110α−/−ΔT mice show normal development of T cell subsets, but slightly reduced numbers of CD4+ T cells in the spleen. “In vitro,” TCR/CD3 plus CD28 activation of naive CD4+ and CD8+ p110α−/−ΔT T cells showed enhanced effector function, particularly IFN-γ secretion, T-bet induction, and Akt, Erk, or P38 activation. Tfh derived from p110α−/−ΔT cells also have enhanced responses when compared to normal mice, and IL-2 expanded p110α−/−ΔT CD8+ T cells had enhanced levels of LAMP-1 and Granzyme B. By contrast, the expansion of p110α−/−ΔT iTreg cells was diminished. Also, p110α−/−ΔT mice had enhanced anti-keyhole limpet hemocyanin (KLH) IFN-γ, or IL-4 responses and IgG1 and IgG2b anti-KLH antibodies, using CFA or Alum as adjuvant, respectively. When compared to WT mice, p110α−/−ΔT mice inoculated with B16.F10 melanoma showed delayed tumor progression. The percentage of CD8+ T lymphocytes was higher and the percentage of Treg cells lower in the spleen of tumor-bearing p110α−/−ΔT mice. Also, IFN-γ production in tumor antigen-activated spleen cells was enhanced. Thus, PI3K p110α plays a significant role in antigen activation and differentiation of CD4+ and CD8+ T lymphocytes modulating antitumor immunity.

Leishmania donovani, the causative agent of Indian visceral leishmaniasis has to face several barriers of the immune system inside the mammalian host for its survival. The complement system is one of the first barriers and consists of a well-balanced network of proteases including S1A family serine proteases (SPs). Inhibitor of serine peptidases (ISPs) is considered as inhibitor of S1A family serine peptidases and is reported to be present in trypanosomes, including Leishmania. In our previous study, we have deciphered the role of ISPs [LdISP1 and L. donovani inhibitor of serine peptidases 2 (LdISP2)] in the survival of L. donovani inside the sandfly midgut. However, the role of theses ISPs in the survival of L. donovani inside mammalian host still remains elusive. In the present study, we have deciphered the inhibitory effect of LdISPs on the host complement S1A serine peptidases, such as C1r/C1s and MASP1/MASP2. Our study suggested that although both rLdISP1 and rLdISP2 inferred strong interaction with C1complex and MBL-associated serine proteases (MASPs) but rLdISP2 showed the stronger inhibitory effect on MASP2 than rLdISP1. Moreover, we found that rLdISP2 significantly reduces the formation of C3, C5 convertase, and membrane attacking complex (MAC) by lectin pathway (LP) resulting in significant reduction in serum mediated lysis of the parasites. The role of LdISP2 on neutrophil elastase-mediated C5aR signaling was also evaluated. Notably, our results showed that infection of macrophages with ISP2-overexpressed Leishmania parasites significantly induces the expression of C5aR both at the transcript and translational level. Simultaneously, infection with ISP2KD parasites results in downregulation of host PI3K/AKT phosphorylation and increased in IL-12 production. Taken together, our findings clearly suggest that LdISP2 promotes parasite survival inside host by inhibiting MAC formation and complement-mediated lysis via LP and by upregulation of C5aR signaling.

Pathogenic gain-of-function mutations in the gene encoding phosphoinositide 3-kinase delta (PI3Kδ) cause activated PI3Kδ syndrome (APDS), a disease characterized by humoral immunodeficiency, lymphadenopathy, and an inability to control persistent viral infections including Epstein–Barr virus (EBV) and cytomegalovirus (CMV) infections. Understanding the mechanisms leading to impaired immune response is important to optimally treat APDS patients. Immunosenescence of CD8+ T cells was suggested to contribute to APDS pathogenesis. However, the constitutive activation of T cells in APDS may also result in T cell exhaustion. Therefore, we studied exhaustion of the CD8+ T cell compartment in APDS patients and compared them with healthy controls and HIV patients, as a control for exhaustion. The subset distribution of the T cell compartment of APDS patients was comparable with HIV patients with decreased naive CD4+ and CD8+ T cells and increased effector CD8+ T cells. Like in HIV+ patients, expression of activation markers and inhibitory receptors CD160, CD244, and programmed death receptor (PD)-1 on CD8+ T cells was increased in APDS patients, indicating exhaustion. EBV-specific CD8+ T cells from APDS patients exhibited an exhausted phenotype that resembled HIV-specific CD8+ T cells in terms of inhibitory receptor expression. Inhibition of PD-1 on EBV-specific CD8+ T cells from APDS patients enhanced in vitro proliferation and effector cytokine production. Based on these results, we conclude that total and EBV-specific CD8+ T cells from APDS patients are characterized by T cell exhaustion. Furthermore, PD-1 checkpoint inhibition may provide a possible therapeutic approach to support the immune system of APDS patients to control EBV and CMV. [ABSTRACT FROM AUTHOR]

Class IA phosphatidylinositol 3-kinase (PI3K) catalytic subunits p110α and p110δ are targets in cancer therapy expressed at high levels in T lymphocytes. The role of p110δ PI3K in normal or pathological immune responses is well established, yet the importance of p110α subunits in T cell-dependent immune responses is not clear. To address this problem, mice with p110α conditionally deleted in CD4+ and CD8+ T lymphocytes (p110α-/-ΔT) were used. p110α-/-ΔT mice show normal development of T cell subsets, but slightly reduced numbers of CD4+ T cells in the spleen. "In vitro," TCR/CD3 plus CD28 activation of naive CD4+ and CD8+ p110α-/-ΔT T cells showed enhanced effector function, particularly IFN-γ secretion, T-bet induction, and Akt, Erk, or P38 activation. Tfh derived from p110α-/-ΔT cells also have enhanced responses when compared to normal mice, and IL-2 expanded p110α-/-ΔT CD8+ T cells had enhanced levels of LAMP-1 and Granzyme B. By contrast, the expansion of p110α-/-ΔT iTreg cells was diminished. Also, p110α-/-ΔT mice had enhanced anti-keyhole limpet hemocyanin (KLH) IFN-γ, or IL-4 responses and IgG1 and IgG2b anti-KLH antibodies, using CFA or Alum as adjuvant, respectively. When compared to WT mice, p110α-/-ΔT mice inoculated with B16.F10 melanoma showed delayed tumor progression. The percentage of CD8+ T lymphocytes was higher and the percentage of Treg cells lower in the spleen of tumor-bearing p110α-/-ΔT mice. Also, IFN-γ production in tumor antigen-activated spleen cells was enhanced. Thus, PI3K p110α plays a significant role in antigen activation and differentiation of CD4+ and CD8+ T lymphocytes modulating antitumor immunity. [ABSTRACT FROM AUTHOR]

Leishmania donovani, the causative agent of Indian visceral leishmaniasis has to face several barriers of the immune system inside the mammalian host for its survival. The complement system is one of the first barriers and consists of a well-balanced network of proteases including S1A family serine proteases (SPs). Inhibitor of serine peptidases (ISPs) is considered as inhibitor of S1A family serine peptidases and is reported to be present in trypanosomes, including Leishmania. In our previous study, we have deciphered the role of ISPs [LdISP1 and L. donovani inhibitor of serine peptidases 2 (LdISP2)] in the survival of L. donovani inside the sandfly midgut. However, the role of theses ISPs in the survival of L. donovani inside mammalian host still remains elusive. In the present study, we have deciphered the inhibitory effect of LdISPs on the host complement S1A serine peptidases, such as C1r/C1s and MASP1/MASP2. Our study suggested that although both rLdISP1 and rLdISP2 inferred strong interaction with C1complex and MBL-associated serine proteases (MASPs) but rLdISP2 showed the stronger inhibitory effect on MASP2 than rLdISP1. Moreover, we found that rLdISP2 significantly reduces the formation of C3, C5 convertase, and membrane attacking complex (MAC) by lectin pathway (LP) resulting in significant reduction in serum mediated lysis of the parasites. The role of LdISP2 on neutrophil elastase-mediated C5aR signaling was also evaluated. Notably, our results showed that infection of macrophages with ISP2-overexpressed Leishmania parasites significantly induces the expression of C5aR both at the transcript and translational level. Simultaneously, infection with ISP2KD parasites results in downregulation of host PI3K/AKT phosphorylation and increased in IL-12 production. Taken together, our findings clearly suggest that LdISP2 promotes parasite survival inside host by inhibiting MAC formation and complement-mediated lysis via LP and by upregulation of C5aR signaling. [ABSTRACT FROM AUTHOR]

The phosphoinositide 3-kinase (PI 3-kinase, PI3K) pathway transduces signals critical for lymphocyte function. PI3K generates the phospholipid PIP3 at the plasma membrane to recruit proteins that contain pleckstrin homology (PH) domains – a conserved domain found in hundreds of mammalian proteins. PH domain–PIP3 interactions allow for rapid signal propagation and confer a spatial component to these signals. The kinases Akt and Itk are key PI3K effectors that bind PIP3 via their PH domains and mediate vital processes – such as survival, activation, and differentiation – in lymphocytes. Here, we review the roles and regulation of PI3K signaling in lymphocytes with a specific emphasis on Akt and Itk. We also discuss these and other PH domain-containing proteins as they relate more broadly to immune cell function. Finally, we highlight the emerging view of PH domains as multifunctional protein domains that often bind both lipid and protein substrates to exert their effects.

Splenic transitional B-cells (T1 and T2) are selected to avoid self-reactivity and to safeguard against autoimmunity, then differentiate into mature follicular (FO-I and FO-II) and marginal zone (MZ) subsets. Transcriptomic analysis by RNA-seq of the five B-cell subsets revealed T1 cell signature genes included RAG suggesting a potential for receptor revision. T1 to T2 B-cell differentiation was marked by a switch from Myb to Myc, increased expression of the PI3K adapter DAP10 and MHC class II. FO-II may be an intermediate in FO-I differentiation and may also become MZ B-cells as suggested by principal component analysis (PCA). MZ B-cells possessed the most distinct transcriptome including downregulation of CD45 phosphatase-associated protein (CD45-AP/PTPRC-AP), as well as upregulation of IL-9R and innate molecules TLR3, TLR7 and bactericidal Perforin-2 (MPEG1). Among the endosomal TLRs, stimulation via TLR3 further enhanced Perforin-2 expression exclusively in MZ B-cells. Using gene-deleted and overexpressing transgenic mice we show that IL-9/IL-9R interaction resulted in rapid activation of STAT1, 3 and 5, primarily in MZ B-cells. Importantly, CD45-AP mutant mice had reduced transitional and increased mature MZ and FO B-cells, suggesting that it prevents premature entry of transitional B-cells to the mature B-cell pool or their survival and proliferation. Together, these findings suggest, developmental plasticity among splenic B-cell subsets, potential for receptor revision in peripheral tolerance whereas enhanced metabolism coincides with T2 to mature B-cell differentiation. Further, unique core transcriptional signatures in MZ B-cells may control their innate features.

The adaptive immune system is equipped to eliminate both tumors and pathogenic microorganisms. It requires a series of complex and coordinated signals to drive the activation, proliferation and differentiation of appropriate T cell subsets. It is now established that changes in cellular activation are coupled to profound changes in cellular metabolism. In addition, emerging evidence now suggest that specific metabolic alterations associated with distinct T cell subsets may be ancillary to their differentiation and influential in their immune functions. The Warburg effect originally used to describe a phenomenon in which most cancer cells relied on aerobic glycolysis for their growth is a key process that sustain T cell activation and differentiation. Here we review how different aspects of metabolism in T cells influence their functions, focusing on the emerging role of key regulators of glucose metabolism such as HIF-1α. A thorough understanding of the role of metabolism in T cell function could provide insights into mechanisms involved in inflammatory-mediated conditions, with the potential for developing novel therapeutic approaches to treat these diseases.

Among numerous cytokines modulating natural killer (NK) cell function, interleukin 15 (IL-15) exerts a broad range of effect from development and homeostasis, to activation of mature NK cells during infection. Its significance is further highlighted by clinical trials in which IL-15 is being used to boost the proliferation and anti-tumor response of NK cells. Among the signal transduction pathways triggered by the engagement of IL-15 receptor with its ligand, the PI3K–AKT–mTOR pathway seems to be critical for the IL-15-mediated activation of NK cells, therefore being responsible for efficient anti-viral and anti-tumor responses. This review provides an overview of the role of IL-15 at multiple stages of NK cell life journey. Understanding the pathway by which IL-15 conveys critical signals for the generation of NK cells with efficient effector functions, in combination with established protocols for NK cell expansion ex vivo, will undoubtedly open new avenues for therapeutic applications for immunomodulation against infections and cancers. [ABSTRACT FROM AUTHOR]

T lymphocytes play a critical role in host defense in all anatomical sites including mucosal surfaces. This not only includes the effector arm of the immune system, but also regulation of immune responses in order to prevent autoimmunity. Genetic targeting of PI3K isoforms suggests that generation of PI(3,4,5)P3 by PI3K plays a critical role in promoting effector T cell responses. Consequently, the 5’- and 3’-inositol poly-phosphatases SHIP1, SHIP2 and PTEN capable of targeting PI(3,4,5)P3 are potential genetic determinants of T cell effector functions in vivo. In addition, the 5’-inositol poly phosphatases SHIP1 and 2 can shunt PI(3,4,5)P3 to the rare but potent signaling phosphoinositide species PI(3,4)P2 and thus these SHIP1/2, and the INPP4A/B enzymes that deplete PI(3,4)P2 may have precise roles in T cell biology to amplify or inhibit effectors of PI3K signaling that are selectively recruited to and activated by PI(3,4)P2. Here we summarize recent genetic and chemical evidence that indicates the inositol poly-phosphatases have important roles in both the effector and regulatory functions of the T cell compartment. In addition, we will discuss future genetic studies that might be undertaken to further elaborate the role of these enzymes in T cell biology as well as potential pharmaceutical manipulation of these enzymes for therapeutic purposes in disease settings where T cell function is a key in vivo target.

Activation of phosphoinositide 3-kinase (PI3K) and Akt (protein kinase B) is a common response triggered by a range of membrane-bound receptors on many cell types. In T lymphocytes, the PI3K-Akt pathway promotes clonal expansion, differentiation, and survival of effector cells and suppresses the generation of regulatory T cells. PI3K activation is tightly controlled by signals through the T cell receptor (TCR) and the co-stimulatory receptor CD28, however sustained and periodic signals from additional co-receptors are now being recognized as critical contributors to the activation of this pathway. Accumulating evidence suggests that many members of the Tumor Necrosis Factor receptor (TNFR) superfamily, TNFR2 (TNFRSF1B), OX40 (TNFRSF4), 4-1BB (TNFRSF9), HVEM (TNFRSF14), and DR3 (TNFRSF25), that are constitutive or inducible on T cells, can directly or indirectly promote activity in the PI3K-Akt pathway. We discuss recent data which suggests that ligation of one TNFR family molecule organizes a signalosome, via TNFR-associated factor (TRAF) adapter proteins in T cell membrane lipid microdomains, that results in the subsequent accumulation of highly concentrated depots of PI3K and Akt in close proximity to TCR signaling units. We propose this may be a generalizable mechanism applicable to other TNFR family molecules that will result in a quantitative contribution of these signalosomes to enhancing and sustaining PI3K and Akt activation triggered by the TCR. We also review data that other TNFR molecules, such as CD40 (TNFRSF5), RANK (TNFRSF11A), FN14 (TNFRSF12A), TACI (TNFRSF13B), BAFFR (TNFRSF13C), and NGFR (TNFRSF16), contribute to the activation of this pathway in diverse cell types through a similar ability to recruit PI3K or Akt into their signaling complexes.