Autoimmune susceptibility gene PTPN2 is required for clearance of adherent-invasive Escherichia coli by integrating bacterial uptake and lysosomal defence.

OBJECTIVES: Alterations in the intestinal microbiota are linked with a wide range of autoimmune and inflammatory conditions, including inflammatory bowel diseases (IBD), where pathobionts penetrate the intestinal barrier and promote inflammatory reactions. In patients with IBD, the ability of intestinal macrophages to efficiently clear invading pathogens is compromised resulting in increased bacterial translocation and excessive immune reactions. Here, we investigated how an IBD-associated loss-of-function variant in the protein tyrosine phosphatase non-receptor type 2 (PTPN2) gene, or loss of PTPN2 expression affected the ability of macrophages to respond to invading bacteria. DESIGN: IBD patient-derived macrophages with wild-type (WT) PTPN2 or carrying the IBD-associated PTPN2 SNP, peritoneal macrophages from WT and constitutive PTPN2-knockout mice, as well as mice specifically lacking PTPN2 in macrophages were infected with non-invasive K12 Escherichia coli, the human adherent-invasive E. coli (AIEC) LF82, or a novel mouse AIEC (mAIEC) strain. RESULTS: Loss of PTPN2 severely compromises the ability of macrophages to clear invading bacteria. Specifically, loss of functional PTPN2 promoted pathobiont invasion/uptake into macrophages and intracellular survival/proliferation by three distinct mechanisms: Increased bacterial uptake was mediated by enhanced expression of carcinoembryonic antigen cellular adhesion molecule (CEACAM)1 and CEACAM6 in PTPN2-deficient cells, while reduced bacterial clearance resulted from defects in autophagy coupled with compromised lysosomal acidification. In vivo, mice lacking PTPN2 in macrophages were more susceptible to mAIEC infection and mAIEC-induced disease. CONCLUSIONS: Our findings reveal a tripartite regulatory mechanism by which PTPN2 preserves macrophage antibacterial function, thus crucially contributing to host defence against invading bacteria.

Programmable Unlocking Nano-Matryoshka-CRISPR Precisely Reverses Immunosuppression to Unleash Cascade Amplified Adaptive Immune Response.

Immune checkpoint blockade (ICB) is an attractive option in cancer therapy, but its efficacy is still less than expected due to the transient and incomplete blocking and the low responsiveness. Herein, an unprecedented programmable unlocking nano-matryoshka-CRISPR system (PUN) targeting programmed cell death ligand 1 (PD-L1) and protein tyrosine phosphatase N2 (PTPN2) is fabricated for permanent and complete and highly responsive immunotherapy. While PUN is inert at normal physiological conditions, enzyme-abundant tumor microenvironment and preternatural intracellular oxidative stress sequentially trigger programmable unlocking of PUN to realize a nano-matryoshka-like release of CRISPR/Cas9. The successful nucleus localization of CRISPR/Cas9 ensures the highly efficient disruption of PD-L1 and PTPN2 to unleash cascade amplified adaptive immune response via revoking the immune checkpoint effect. PD-L1 downregulation in tumor cells not only disrupts PD-1/PD-L1 interaction to attenuate the immunosurveillance evasion but also spurs potent immune T cell responses to enhance adaptive immunity. Synchronously, inhibition of JAK/STAT pathway is relieved by deleting PTPN2, which promotes tumor susceptibility to CD8+ T cells depending on IFN-γ, thus further amplifying adaptive immune responses. Combining these advances together, PUN exhibits optimal antitumor efficiency and long-term immune memory with negligible toxicity, which provides a promising alternative to current ICB therapy.

Protein tyrosine phosphatase nonreceptor type 2 controls colorectal cancer development.

Protein tyrosine phosphatase nonreceptor type 2 (PTPN2) recently emerged as a promising cancer immunotherapy target. We set out to investigate the functional role of PTPN2 in the pathogenesis of human colorectal carcinoma (CRC), as its role in immune-silent solid tumors is poorly understood. We demonstrate that in human CRC, increased PTPN2 expression and activity correlated with disease progression and decreased immune responses in tumor tissues. In particular, stage II and III tumors displayed enhanced PTPN2 protein expression in tumor-infiltrating T cells, and increased PTPN2 levels negatively correlated with expression of PD-1, CTLA4, STAT1, and granzyme A. In vivo, T cell- and DC-specific PTPN2 deletion reduced tumor burden in several CRC models by promoting CD44+ effector/memory T cells, as well as CD8+ T cell infiltration and cytotoxicity in the tumor. In direct relevance to CRC treatment, T cell-specific PTPN2 deletion potentiated anti-PD-1 efficacy and induced antitumor memory formation upon tumor rechallenge in vivo. Our data suggest a role for PTPN2 in suppressing antitumor immunity and promoting tumor development in patients with CRC. Our in vivo results identify PTPN2 as a key player in controlling the immunogenicity of CRC, with the strong potential to be exploited for cancer immunotherapy.

PTPN2 negatively regulates macrophage inflammation in atherosclerosis.

Atherosclerosis is the main cause of cardiovascular disease. Systemic inflammation is one important characteristic in atherosclerosis. Pro-inflammatory macrophages can secrete inflammatory factors and promote the inflammation of atherosclerosis. It has a great value for the treatment of atherosclerosis by inhibiting the release of inflammatory factors in macrophages. However, the detailed mechanism of this process is still unclear. In this study, we constructed an APOE-/- mice model of atherosclerosis to research the molecular mechanism of atherosclerosis. Protein tyrosine phosphatase non-receptor type 2 (PTPN2), an anti-inflammatory gene, was dramatically decreased in inflammatory mice. Deletion of PTPN2 could significantly induce monocytes toward M1 phenotype of macrophages, enhance the secretion of IL-12 and IL-1, and promote cell proliferation, invasion and metastasis. Mechanism research showed that PTPN2-mediated p65/p38/STAT3 de-phosphorylation could block the process of macrophage inflammation. In vivo experiments showed that PTPN2 may effectively inhibit the inflammatory response during atherosclerosis. In conclusion, we uncovered the negative role of PTPN2 in the occurrence of atherosclerosis, and this study provides a new potential target for atherosclerosis treatment.

A CRISPR-Cas9 delivery system for in vivo screening of genes in the immune system.

Therapies that target the function of immune cells have significant clinical efficacy in diseases such as cancer and autoimmunity. Although functional genomics has accelerated therapeutic target discovery in cancer, its use in primary immune cells is limited because vector delivery is inefficient and can perturb cell states. Here we describe CHIME: CHimeric IMmune Editing, a CRISPR-Cas9 bone marrow delivery system to rapidly evaluate gene function in innate and adaptive immune cells in vivo without ex vivo manipulation of these mature lineages. This approach enables efficient deletion of genes of interest in major immune lineages without altering their development or function. We use this approach to perform an in vivo pooled genetic screen and identify Ptpn2 as a negative regulator of CD8+ T cell-mediated responses to LCMV Clone 13 viral infection. These findings indicate that this genetic platform can enable rapid target discovery through pooled screening in immune cells in vivo.

Reduced expression of phosphatase PTPN2 promotes pathogenic conversion of Tregs in autoimmunity.

Genetic variants at the PTPN2 locus, which encodes the tyrosine phosphatase PTPN2, cause reduced gene expression and are linked to rheumatoid arthritis (RA) and other autoimmune diseases. PTPN2 inhibits signaling through the T cell and cytokine receptors, and loss of PTPN2 promotes T cell expansion and CD4- and CD8-driven autoimmunity. However, it remains unknown whether loss of PTPN2 in FoxP3+ regulatory T cells (Tregs) plays a role in autoimmunity. Here we aimed to model human autoimmune-predisposing PTPN2 variants, the presence of which results in a partial loss of PTPN2 expression, in mouse models of RA. We identified that reduced expression of Ptpn2 enhanced the severity of autoimmune arthritis in the T cell-dependent SKG mouse model and demonstrated that this phenotype was mediated through a Treg-intrinsic mechanism. Mechanistically, we found that through dephosphorylation of STAT3, PTPN2 inhibits IL-6-driven pathogenic loss of FoxP3 after Tregs have acquired RORγt expression, at a stage when chromatin accessibility for STAT3-targeted IL-17-associated transcription factors is maximized. We conclude that PTPN2 promotes FoxP3 stability in mouse RORγt+ Tregs and that loss of function of PTPN2 in Tregs contributes to the association between PTPN2 and autoimmunity.

TC-PTP and PTP1B: Regulating JAK-STAT signaling, controlling lymphoid malignancies.

Lymphoid malignancies are characterized by an accumulation of genetic lesions that act co-operatively to perturb signaling pathways and alter gene expression programs. The Janus kinases (JAK)-signal transducers and activators of transcription (STATs) pathway is one such pathway that is frequently mutated in leukemia and lymphoma. In response to cytokines and growth factors, a cascade of reversible tyrosine phosphorylation events propagates the JAK-STAT pathway from the cell surface to the nucleus. Activated STAT family members then play a fundamental role in establishing the transcriptional landscape of the cell. In leukemia and lymphoma, somatic mutations have been identified in JAK and STAT family members, as well as, negative regulators of the pathway. Most recently, inactivating mutations in the protein tyrosine phosphatase (PTP) genes PTPN1 (PTP1B) and PTPN2 (TC-PTP) were sequenced in B cell lymphoma and T cell acute lymphoblastic leukemia (T-ALL) respectively. The loss of PTP1B and TC-PTP phosphatase activity is associated with an increase in cytokine sensitivity, elevated JAK-STAT signaling, and changes in gene expression. As inactivation mutations in PTPN1 and PTPN2 are restricted to distinct subsets of leukemia and lymphoma, a future challenge will be to identify in which cellular contexts do they contributing to the initiation or maintenance of leukemogenesis or lymphomagenesis. As well, the molecular mechanisms by which PTP1B and TC-PTP loss co-operates with other genetic aberrations will need to be elucidated to design more effective therapeutic strategies.

Non-HLA gene effects on the disease process of type 1 diabetes: From HLA susceptibility to overt disease.

In addition to the HLA region numerous other gene loci have shown association with type 1 diabetes. How these polymorphisms exert their function has not been comprehensively described, however. We assessed the effect of 39 single nucleotide polymorphisms (SNP) on the development of autoantibody positivity, on progression from autoantibody positivity to clinical disease and on the specificity of the antibody initiating the autoimmune process in 521 autoantibody-positive and 989 control children from a follow-up study starting from birth. Interestingly, PTPN2 rs45450798 gene polymorphism was observed to strongly affect the progression rate of beta-cell destruction after the appearance of humoral beta-cell autoimmunity. Moreover, primary autoantigen dependent associations were also observed as effect of the IKZF4-ERBB3 region on the progression rate of ß-cell destruction was restricted to children with GAD antibodies as their first autoantibody whereas the effect of the INS rs 689 polymorphism was observed among subjects with insulin as the primary autoantigen. In the whole study cohort, INS rs689, PTPN22 rs2476601 and IFIH1 rs1990760 polymorphisms were associated with the appearance of beta-cell autoantibodies. These findings provide new insights into the role of genetic factors implicated in the pathogenesis of type 1 diabetes. The effect of some of the gene variants is restricted to control the initiation of ß-cell autoimmunity whereas others modify the destruction rate of the ß-cells. Furthermore, signs of primary autoantigen-related pathways were detected.

PTPN2 restrains CD8⁺ T cell responses after antigen cross-presentation for the maintenance of peripheral tolerance in mice.

Antigen cross-presentation by dendritic cells is crucial for priming cytotoxic CD8(+) T cells to invading pathogens and tumour antigens, as well as mediating peripheral tolerance to self-antigens. The protein tyrosine phosphatase N2 (PTPN2) attenuates T cell receptor (TCR) signalling and tunes CD8(+) T cell responses in vivo. In this study we have examined the role of PTPN2 in the maintenance of peripheral tolerance after the cross-presentation of pancreatic ß-cell antigens. The transfer of OVA-specific OT-I CD8(+) T cells (C57BL/6) into RIP-mOVA recipients expressing OVA in pancreatic ß-cells only results in islet destruction when OVA-specific CD4(+) T cells are co-transferred. Herein we report that PTPN2-deficient OT-I CD8(+) T cells transferred into RIP-mOVA recipients acquire CTL activity and result in ß cell destruction and the development of diabetes in the absence of CD4(+) help. These studies identify PTPN2 as a critical mediator of peripheral T cell tolerance limiting CD8(+) T cell responses after the cross-presentation of self-antigens. Our findings reveal a mechanism by which PTPN2 SNPs might convert a tolerogenic CD8(+) T cell response into one capable of causing the destruction of pancreatic ß-cells. Moreover, our results provide insight into potential approaches for enhancing T cell-mediated immunity and/or T cell adoptive tumour immunotherapy.

Protein tyrosine phosphatases in lymphocyte activation and autoimmunity.

Lymphocyte activation must be tightly regulated to ensure sufficient immunity to pathogens and prevent autoimmunity. Protein tyrosine phosphatases (PTPs) serve critical roles in this regulation by controlling the functions of key receptors and intracellular signaling molecules in lymphocytes. In some cases, PTPs inhibit lymphocyte activation, whereas in others they promote it. Here we discuss recent progress in elucidating the roles and mechanisms of action of PTPs in lymphocyte activation. We also review the accumulating evidence that genetic alterations in PTPs are involved in human autoimmunity.

PTPN2 but not PTPN22 is associated with Crohn's disease in a New Zealand population.

Recent genome-wide association studies have provided evidence for the involvement of the genes PTPN2 and PTPN22 in the pathogenesis of Crohn's disease (CD). We investigated whether genetic variants in these genes were associated with CD in a New Zealand population. Single-nucleotide polymorphisms (SNPs) rs2542151 (PTPN2) and rs2476601 (PTPN22) were genotyped in 315 CD cases and 481 controls. In this sample, we were able to confirm an association between CD and PTPN2 (genotypic P = 0.019 and allelic P = 0.011), and phenotypic analysis showed an association of this SNP with late age at first diagnosis, inflammatory and penetrating CD behaviour, requirement of bowel resection and being a smoker at diagnosis. There was no evidence for an association with PTPN22.

The role of protein tyrosine phosphatases in the regulation of allergic asthma: implication of TC-PTP and PTP-1B in the modulation of disease development.

Protein tyrosine phosphorylation is an important early event in the signal transduction of numerous cell receptors involved in the immune response. The implication of protein tyrosine kinases in allergic asthma is well recognized, but the role of protein tyrosine phosphatases (PTPs) remains poorly understood. However, we recently reported that global inhibition of PTPs during either the allergen-sensitization phase or the allergen-challenge phase reduced the development of asthma and that this correlated with an increased T helper 1 (Th1) response in both lung and spleen tissues. Therefore, in this study we investigated individual roles of PTPs involved in regulating the immune response. We observed that genetic deficiency for PTP-1B resulted in increased recruitment of lung inflammatory cells, while protein tyrosine phosphatase-phosphatase and tensin homologue deleted (PTP-PEST)-deficient mice exhibited a phenotype similar to that of wild-type mice. Importantly, we found that a heterozygous mutation of T cell PTP (TC-PTP) dramatically abrogates immunoglobulin E production and reduces the recruitment of inflammatory cells to the lung, conferring an important role for TC-PTP in the development of allergic asthma. As opposed to other studies on Src homology phosphatase-1 (SHP-1) deficiency, specific acute SHP-1 inhibition during allergen challenge did not affect disease outcome. Collectively, our results underscore the importance of PTPs in the development of allergic asthma.

T-cell protein tyrosine phosphatase is a key regulator in immune cell signaling: lessons from the knockout mouse model and implications in human disease.

The immune system requires for its proper ontogeny, differentiation, and maintenance the function of several tyrosine kinases and adapters that create and modify tyrosine phosphorylation sites. Tyrosine phosphorylation is a crucial protein modification in immune cell signaling and can be reversed by protein tyrosine phosphatases (PTPs). Much progress has been made in identifying and understanding PTP function in the immune system. In this review, we present one of these proteins, named T-cell PTPs (TC-PTP) (gene name PTPN2), a classical, non-receptor PTP that is ubiquitously expressed with particularly high expression in hematopoietic tissues. TC-PTP is remarkable not only by the fact that it appears to influence most, if not all, cells involved in the development of the immune system, from stem cells to differentiated lineages, but also recent findings have positioned it at the core of several human diseases from autoimmune disease to cancer.