Protein tyrosine phosphatase, non-receptor type 22 (PTPN22) regulates a panoply of leukocyte signaling pathways. A single nucleotide polymorphism (SNP) in
,
, is associated with increased risk of ...Type 1 Diabetes (T1D) and other autoimmune diseases. Over the past decade PTPN22 has been studied intensely in T cell receptor (TCR) and B cell receptor (BCR) signaling. However, the effect of the minor allele on PTPN22 function in TCR signaling is controversial with some reports concluding it has enhanced function and blunts TCR signaling and others reporting it has reduced function and increases TCR signaling. More recently, the core function of PTPN22 as well as functional derangements imparted by the autoimmunity-associated variant allele of PTPN22 have been examined in monocytes, macrophages, dendritic cells, and neutrophils. In this review we will discuss the known functions of PTPN22 in human cells, and we will elaborate on how autoimmunity-associated variants influence these functions across the panoply of immune cells that express PTPN22. Further, we consider currently unresolved questions that require clarification on the role of PTPN22 in immune cell function.
Prior studies identified HLA class-II and 57 additional loci as contributors to genetic susceptibility for type 1 diabetes (T1D). We hypothesized that race and/or ethnicity would be contextually ...important for evaluating genetic risk markers previously identified from Caucasian/European cohorts. We determined the capacity for a combined genetic risk score (GRS) to discriminate disease-risk subgroups in a racially and ethnically diverse cohort from the southeastern U.S. including 637 T1D patients, 46 at-risk relatives having two or more T1D-related autoantibodies (≥2AAb
), 790 first-degree relatives (≤1AAb
), 68 second-degree relatives (≤1 AAb
), and 405 controls. GRS was higher among Caucasian T1D and at-risk subjects versus ≤ 1AAb
relatives or controls (P < 0.001). GRS receiver operating characteristic AUC (AUROC) for T1D versus controls was 0.86 (P < 0.001, specificity = 73.9%, sensitivity = 83.3%) among all Caucasian subjects and 0.90 for Hispanic Caucasians (P < 0.001, specificity = 86.5%, sensitivity = 84.4%). Age-at-diagnosis negatively correlated with GRS (P < 0.001) and associated with HLA-DR3/DR4 diplotype. Conversely, GRS was less robust (AUROC = 0.75) and did not correlate with age-of-diagnosis for African Americans. Our findings confirm GRS should be further used in Caucasian populations to assign T1D risk for clinical trials designed for biomarker identification and development of personalized treatment strategies. We also highlight the need to develop a GRS model that accommodates racial diversity.
The IFN‐stimulated gene ubiquitin‐specific proteinase 18 (USP18) encodes a protein that negatively regulates T1 IFN signaling via stearic inhibition of JAK1 recruitment to the IFN‐α receptor 2 ...subunit (IFNAR2). Here, we demonstrate that USP18 expression is induced by HIV‐1 in a T1 IFN‐dependent manner. Experimental depletion of USP18 by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9) gene editing results in a significant restriction of HIV‐1 replication in an induced pluripotent stem cell (iPSC)‐derived macrophage model. In the absence of USP18, macrophages have increased responsiveness to stimulation with T1 IFNs with prolonged phosphorylation of STAT1 and STAT2 and increased expression of IFN‐stimulated genes that are key for antiviral responses. Interestingly, HIV‐1 requires some signaling through the T1 IFN receptor to replicate efficiently because a neutralizing antibody that inhibits T1 IFN activity reduces HIV‐1 replication rate in monocyte‐derived macrophages. USP18 induction by HIV‐1 tunes the IFN response to optimal levels allowing for efficient transcription from the HIV‐1 LTR promoter while minimizing the T1 IFN‐induced antiviral response that would otherwise restrict viral replication and spread. Finally, iPSC and CRISPR/Cas9 gene targeting offer a powerful tool to study host factors that regulate innate immune responses.
USP18 promotes HIV‐1 replication by negatively regulating expression of antiviral genes induced by T1 IFN signaling.
Adeno-associated virus (AAV) is a replication-deficient parvovirus that is extensively used as a gene therapy vector. CD8+ T-cell responses against the AAV capsid protein can, however, affect ...therapeutic efficacy. Little is known about the in vivo mechanism that leads to the crosspriming of CD8+ T cells against the input viral capsid antigen. In this study, we report that the Toll-like receptor 9 (TLR9)–MyD88 pattern-recognition receptor pathway is uniquely capable of initiating this response. By contrast, the absence of TLR2, STING, or the addition of TLR4 agonist has no effect. Surprisingly, both conventional dendritic cells (cDCs) and plasmacytoid DCs (pDCs) are required for the crosspriming of capsid-specific CD8+ T cells, whereas other antigen-presenting cells are not involved. TLR9 signaling is specifically essential in pDCs but not in cDCs, indicating that sensing of the viral genome by pDCs activates cDCs in trans to cross-present capsid antigen during CD8+ T-cell activation. Cross-presentation and crosspriming depend not only on TLR9, but also on interferon type I signaling, and both mechanisms can be inhibited by administering specific molecules to prevent induction of capsid-specific CD8+ T cells. Thus, these outcomes directly point to therapeutic interventions and demonstrate that innate immune blockade can eliminate unwanted immune responses in gene therapy.
•Crosspriming of AAV capsid-specific CD8+ T cells requires cooperation between distinct subsets of DCs.•Innate immune sensing of the viral DNA genome induces cross-presentation of viral capsid in trans.
Adeno-associated virus (AAV) vectors are widely used in clinical gene therapy to correct genetic disease by in vivo gene transfer. Although the vectors are useful, in part because of their limited ...immunogenicity, immune responses directed at vector components have complicated applications in humans. These include, for instance, innate immune sensing of vector components by plasmacytoid dendritic cells (pDCs), which sense the vector DNA genome via Toll-like receptor 9. Adaptive immune responses employ antigen presentation by conventional dendritic cells (cDCs), which leads to cross-priming of capsid-specific CD8+ T cells. In this study, we sought to determine the mechanisms that promote licensing of cDCs, which is requisite for CD8+ T cell activation. Blockage of type 1 interferon (T1 IFN) signaling by monoclonal antibody therapy prevented cross-priming. Furthermore, experiments in cell-type-restricted knockout mice showed a specific requirement for the receptor for T1 IFN (IFNaR) in cDCs. In contrast, natural killer (NK) cells are not needed, indicating a direct rather than indirect effect of T1 IFN on cDCs. In addition, co-stimulation by CD4+ T cells via CD40-CD40L was required for cross-priming, and blockage of co-stimulation but not of T1 IFN additionally reduced antibody formation against capsid. These mechanistic insights inform the development of targeted immune interventions.
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Immune responses complicate the use of adeno-associated virus (AAV) vectors in human gene therapy. Shirley et al. define a mechanism by which cross-presentation of viral capsid results in activation of CD8+ T cells, which involves sensing of IFN I by conventional dendritic cells and co-stimulation by CD4+ T helper cells.
Sepsis, the systemic inflammatory response to microbial infection, induces changes in both innate and adaptive immunity that presumably lead to increased susceptibility to secondary infections, ...multiorgan failure, and death. Using a model of murine polymicrobial sepsis whose severity approximates human sepsis, we examined outcomes and defined requirements for survival after secondary Pseudomonas aeruginosa pneumonia or disseminated Listeria monocytogenes infection. We demonstrate that early after sepsis neutrophil numbers and function are decreased, whereas monocyte recruitment through the CCR2/MCP-1 pathway and function are enhanced. Consequently, lethality to Pseudomonas pneumonia is increased early but not late after induction of sepsis. In contrast, lethality to listeriosis, whose eradication is dependent upon monocyte/macrophage phagocytosis, is actually decreased both early and late after sepsis. Adaptive immunity plays little role in these secondary infectious responses. This study demonstrates that sepsis promotes selective early, impaired innate immune responses, primarily in neutrophils, that lead to a pathogen-specific, increased susceptibility to secondary infections.
HIV-1 replication and microbial translocation occur concomitant with systemic immune activation. This study delineates mechanisms of immune activation and CD4 T-cell decline in pediatric HIV-1 ...infection.
Cross-sectional and longitudinal cellular and soluble plasma markers for inflammation were evaluated in 14 healthy and 33 perinatally HIV-1-infected pediatric study volunteers prior to and over 96 weeks of protease-inhibitor-containing combination antiretroviral therapy (ART). All HIV-1-infected patients reconstituted CD4 T cells either with suppression of viremia or rebound of drug-resistant virus.
Systemic immune activation was determined by polychromatic flow cytometry of blood lymphocytes and ELISA for plasma soluble CD27, soluble CD14, and tumor necrosis factor. Microbial translocation was evaluated by limulus amebocyte lysate assay to detect bacterial lipopolysaccharide (LPS) and ELISA for antiendotoxin core antigen immunoglobulin M (IgM) antibodies. Immune activation markers were compared with viral load, CD4 cell percentage, and LPS by regression models. Comparisons between healthy and HIV-1-infected or between different viral outcome groups were performed by nonparametric rank sum.
Microbial translocation was detected in healthy infants but resolved with age (P < 0.05). LPS and soluble CD14 levels were elevated in all HIV-1-infected patients (P < 0.05 and P < 0.0001, respectively) and persisted even if CD4 T cells were fully reconstituted, virus optimally suppressed, and lymphocyte activation resolved by ART. Children with CD4 T-cell reconstitution but viral rebound following ART continued to display high levels of soluble CD27.
Microbial translocation in pediatric HIV-1 infection is associated with persistent monocyte/macrophage activation independent of viral replication or T-cell activation.
Stem cell-derived β-like cells (sBC) carry the promise of providing an abundant source of insulin-producing cells for use in cell replacement therapy for patients with diabetes, potentially allowing ...widespread implementation of a practical cure. To achieve their clinical promise, sBC need to function comparably with mature adult β-cells, but as yet they display varying degrees of maturity. Indeed, detailed knowledge of the events resulting in human β-cell maturation remains obscure. Here we show that sBC spontaneously self-enrich into discreet islet-like cap structures within in vitro cultures, independent of exogenous maturation conditions. Multiple complementary assays demonstrate that this process is accompanied by functional maturation of the self-enriched sBC (seBC); however, the seBC still contain distinct subpopulations displaying different maturation levels. Interestingly, the surface protein ENTPD3 (also known as nucleoside triphosphate diphosphohydrolase-3 NDPTase3) is a specific marker of the most mature seBC population and can be used for mature seBC identification and sorting. Our results illuminate critical aspects of in vitro sBC maturation and provide important insights toward developing functionally mature sBC for diabetes cell replacement therapy.
Reactive oxygen species (ROS) are critical in driving the onset of type 1 diabetes (T1D). Ablation of ROS derived from phagocytic NADPH oxidase 2 is protective against autoimmune diabetes in ...non-obese diabetic (NOD
mice. However, the mechanisms of NADPH oxidase 2-derived ROS in T1D pathogenesis need to be elucidated. Here, we have examined the role of
(the regulatory subunit of NADPH oxidase 2) in dendritic cells (DC).
-mutant DCs exhibit reduced ability to activate autoreactive CD8
T cells despite no difference in co-stimulatory molecule expression or pro-inflammatory cytokine production. When provided with exogenous whole-protein antigen,
-mutant NOD DCs showed strong phagosome acidification and rapid antigen degradation, which lead to an absence of protein translocation into the cytoplasm and deficient antigenic peptide loading on MHC Class I molecules.
This study demonstrates that Ncf1 (p47
) is required for activation and effector function of CD8
T cells by acting both intrinsically within the T cell as well as within professional antigen presenting cells.
ROS promote CD8
T cell activation by facilitating autoantigen cross-presentation by DCs. ROS scavengers could potentially represent an important component of therapies aiming to disrupt autoantigen presentation and activation of CD8
T cells in individuals at-risk for developing T1D.
Type 1 diabetes (T1D) is a disease that arises due to complex immunogenetic mechanisms. Key cell-cell interactions involved in the pathogenesis of T1D are activation of autoreactive T cells by ...dendritic cells (DC), migration of T cells across endothelial cells (EC) lining capillary walls into the islets of Langerhans, interaction of T cells with macrophages in the islets, and killing of β-cells by autoreactive CD8
T cells. Overall, pathogenic cell-cell interactions are likely regulated by the individual's collection of genetic T1D-risk variants. To accurately model the role of genetics, it is essential to build systems to interrogate single candidate genes in isolation during the interactions of cells that are essential for disease development. However, obtaining single-donor matched cells relevant to T1D is a challenge. Sourcing these genetic variants from human induced pluripotent stem cells (iPSC) avoids this limitation. Herein, we have differentiated iPSC from one donor into DC, macrophages, EC, and β-cells. Additionally, we also engineered T cell avatars from the same donor to provide an
platform to study genetic influences on these critical cellular interactions. This proof of concept demonstrates the ability to derive an isogenic system from a single donor to study these relevant cell-cell interactions. Our system constitutes an interdisciplinary approach with a controlled environment that provides a proof-of-concept for future studies to determine the role of disease alleles (e.g.
in regulating cell-cell interactions and cell-specific contributions to the pathogenesis of T1D.
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