Protein Tyrosine Phosphatase Non-receptor Type Research Articles

Erythropoietin inhibits apoptosis of retinal ganglion cells induced by high glucose through JNK signaling pathway

The aim of this study was to investigate the effect of erythropoietin (EPO) on the apoptosis of retinal ganglion cells (RGCs) induced by high glucose and its mechanism. Rat primary RGCs were extracted to establish high glucose-induced apoptosis models using a 30 mM high-glucose medium. Then flow cytometry, cell counting kit-8 (CCK-8) assay and Western blotting assay were performed to detect the effects of high-, medium- and low-dose EPO on the apoptosis of RGCs induced by high glucose. Next, the molecular mechanism by which EPO suppressed the high glucose-induced apoptosis of RGCs was explored via gene array assay and bioinformatics analysis. The results and mechanism of bioinformatics analysis were verified by Western blotting assay. Finally, the small interfering ribonucleic acid (siRNA) experiment was applied to knock down tyrosine-protein phosphatase non-receptor type 1 (PTPN1) and PTPN11 to verify their roles in the inhibition of EPO on the apoptosis of RGCs triggered by high glucose. Flow cytometry-Annexin V/propidium iodide (PI) staining and CCK-8 assay confirmed that the high-, medium- and low-dose EPO inhibited the apoptosis of RGCs induced by high glucose in a dose-dependent manner (P<0.05). Subsequently, Western blotting assay results manifested that the high-, medium- and low-dose EPO reduced the expression levels of apoptosis-related proteins active-cysteinyl aspartate specific proteinase 3 (Caspase 3) and active- Caspase 9 in a dose-dependent manner (P<0.05). Moreover, according to gene array assay and bioinformatics analysis results, the c-Jun N-terminal kinase (JNK) signaling pathway, PTPN1 and PTPN11 might exert crucial effects in the inhibition of EPO on the apoptosis of RGCs induced by high glucose. Western blotting assay results also demonstrated that, compared with the high-glucose treatment, the high-dose EPO treatment decreased the protein expression level of phosphorylated (p)-JNK1/JNK but increased the protein expression levels of PTPN1 and PTPN11 (P<0.05). Moreover, flow cytometry-Annexin V/PI staining and CCK-8 assay results revealed that in EPO-treated cells, knocking down PTPN1 and PTPN11 significantly reversed the protective effect of EPO against high glucose-induced retinal ganglion cell apoptosis (P<0.05). Lastly, Western blotting assay illustrated that knocking down PTPN1 and PTPN11 significantly abolished the inhibition of high-dose EPO on the JNK signaling pathway. EPO may suppress the JNK signaling pathway by raising the expression levels of PTPN1 and PTPN11, so as to inhibit the apoptosis of RGCs triggered by high glucose.

Type 1 Diabetes and Autoimmune Thyroid Disease-The Genetic Link.

Type 1 diabetes (T1D) and autoimmune thyroid disease (AITD) are the most frequent chronic autoimmune diseases worldwide. Several autoimmune endocrine and non-endocrine disorders tend to occur together. T1D and AITD often cluster in individuals and families, seen in the formation of autoimmune polyendocrinopathy (AP). The close relationship between these two diseases is largely explained by sharing a common genetic background. The HLA antigens DQ2 (DQA1*0501-DQB1*0201) and DQ8 (DQA1*0301-DQB1*0302), tightly linked with DR3 and DR4, are the major common genetic predisposition. Moreover, functional single nucleotide polymorphisms (or rare variants) of various genes, such as the cytotoxic T-lymphocyte- associated antigen (CTLA4), the protein tyrosine phosphatase non-receptor type 22 (PTPN22), the interleukin-2 Receptor (IL2Ra), the Vitamin D receptor (VDR), and the tumor-necrosis-factor-α (TNF) that are involved in immune regulation have been identified to confer susceptibility to both T1D and AITD. Other genes including cluster of differentiation 40 (CD40), the forkhead box P3 (FOXP3), the MHC Class I Polypeptide-Related Sequence A (MICA), insulin variable number of tandem repeats (INS-VNTR), the C-Type Lectin Domain Containing 16A (CLEC16A), the Erb-B2 Receptor Tyrosine Kinase 3 (ERBB3) gene, the interferon-induced helicase C domain-containing protein 1 (IFIH1), and various cytokine genes are also under suspicion to increase susceptibility to T1D and AITD. Further, BTB domain and CNC homolog 2 (BACH2), C-C motif chemokine receptor 5 (CCR5), SH2B adaptor protein 3 (SH2B3), and Rac family small GTPase 2 (RAC2) are found to be associated with T1D and AITD by various independent genome wide association studies and overlap in our list, indicating a strong common genetic link for T1D and AITD. As several susceptibility genes and environmental factors contribute to the disease aetiology of both T1D and AITD and/or AP subtype III variant (T1D+AITD) simultaneously, all patients with T1D should be screened for AITD, and vice versa.

PTPN14 deficiency alleviates podocyte injury through suppressing inflammation and fibrosis by targeting TRIP6 in diabetic nephropathy

Diabetic nephropathy (DN) is a common complication of diabetes, and a leading cause of end-stage renal disease. However, the pathogenesis that contributes to DKD is still not fully understood. Protein tyrosine phosphatase non-receptor type 14 (PTPN14), a non receptor tyrosine phosphatase, has numerous cellular events, such as inflammation and cell death. But its potential on DKD has not been investigated yet. In this study, we found that PTPN14 expression was markedly up-regulated in kidney samples of DKD patients, which were confirmed in diabetic mice and were clearly localized in glomeruli. The diabetic mouse model was established using streptozotocin (STZ) in wild type (WT) or PTPN knockout (KO) mice. After, STZ challenge, STZ mice displayed improved kidney functions. The results also showed that STZ-induced histological changes and podocyte injury in renal tissues, which were effectively alleviated by PTPN14 deletion. Moreover, PTPN14 deficiency significantly mitigated inflammatory response and fibrosis in glomeruli of STZ-challenged mice through restraining the activation of nuclear factor-κB (NF-κB) and transforming growth factor (TGF)-β1 signaling pathways, respectively. The inhibitory effects of PTPN14 suppression on inflammation and fibrosis were confirmed in high glucose (HG)-incubated podocytes. We further found that thyroid receptor interactor protein 6 (TRIP6) expression was dramatically up-regulated in glomeruli of STZ-challenged mice, and was abolished by PTPN14 deletion, which was confirmed in HG-treated podocytes with PTPN14 knockdown. Intriguingly, our in vitro studies showed that PTPN14 directly interacted with TRIP6. Of note, over-expressing TRIP6 markedly abrogated the effects of PTPN14 silence to restrict inflammatory response and fibrosis in HG-incubated podocytes. Taken together, our findings demonstrated that targeting PTPN14 may provide feasible therapies for DKD treatment.

Influence of PTPN22 Allotypes on Innate and Adaptive Immune Function in Health and Disease.

Protein tyrosine phosphatase, non-receptor type 22 (PTPN22) regulates a panoply of leukocyte signaling pathways. A single nucleotide polymorphism (SNP) in PTPN22, rs2476601, 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.

Association of PTPN22-C1858T Polymorphism With Susceptibility to Mycobacterium tuberculosis and Mycobacterium leprae Infection: A Meta-Analysis

It was previously published that single-nucleotide polymorphism rs2476601 (PTPN22 [protein tyrosine phosphatase non-receptor type 22]-C1858T) might be related to increased sensibility to Mycobacterium tuberculosis and M. leprae infection. However, the results were inconclusive despite a high degree of similarity between both parameters. Herein, we carried out this meta-analysis to systematically summarize and articulate the correlation between PTPN22-C1858T polymorphism and mycobacterial infection. The susceptibility of PTPN22-C1858T carriers with autoimmune conditions receiving immunosuppressive therapy to M. tuberculosis and M. leprae infection was determined. A systematic retrieval of studies on relevance of PTPN22-C1858T polymorphism to susceptibility of M. tuberculosis or M. leprae infection was performed in Chinese National Knowledge Infrastructure, PubMed and Embase databases. We regarded Odds ratios (ORs) and 95% confidence intervals (CIs) as the determined effect size. Finally, four and two case-control studies on tuberculosis and leprosy, respectively, were included. In all genetic models, without indicated association between PTPN22-C1858T polymorphism and tuberculosis’s susceptibility. [C versus T: OR = 0.22 (95% CI: 0.09–0.50, PH = 0.887); CT versus CC: OR = 0.21 (95% CI: 0.09–0.49, PH = 0.889); TT+CT versus CC: OR = 0.21 (95% CI: 0.09–0.49, PH = 0.889)]. A significantly increased risk of leprosy was perceived in patients with the PTPN22-C1858T polymorphism [C versus T: OR = 2.82 (95% CI: 1.02–7.81, PH = 0.108)]. While the PTPN22-C1858T polymorphism is irrelevant to higher susceptibility to the infection of M. tuberculosis in Caucasians and Asians, it is relevant to increased susceptibility to the infection of M. leprae. However, the results of M. leprae are supposed to interpreted with prudence owing to the limited quantity of studies and heterogeneity. Further well-designed studies with sufficient populations are required to verify our conclusions.

Proteasome regulation by reversible tyrosine phosphorylation at the membrane.

Reversible phosphorylation has emerged as an important mechanism for regulating 26S proteasome function in health and disease. Over 100 phospho-tyrosine sites of the human proteasome have been detected, and yet their function and regulation remain poorly understood. Here we show that the 19S subunit Rpt2 is phosphorylated at Tyr439, a strictly conserved residue within the C-terminal HbYX motif of Rpt2 that is essential for 26S proteasome assembly. Unexpectedly, we found that Y439 phosphorylation depends on Rpt2 membrane localization mediated by its N-myristoylation. Multiple receptors tyrosine kinases can trigger Rpt2-Y439 phosphorylation by activating Src, a N-myristoylated tyrosine kinase. Src directly phosphorylates Rpt2-Y439 in vitro and negatively regulates 26S proteasome activity at cellular membranes, which can be reversed by the membrane-associated isoform of protein tyrosine phosphatase nonreceptor type 2 (PTPN2). In H1975 lung cancer cells with activated Src, blocking Rpt2-Y439 phosphorylation by the Y439F mutation conferred partial resistance to the Src inhibitor saracatinib both in vitro and in a mouse xenograft tumor model, and caused significant changes of cellular responses to saracatinib at the proteome level. Our study has defined a novel mechanism involved in the spatial regulation of proteasome function and provided new insights into tyrosine kinase inhibitor-based anticancer therapies.

Harmine alleviates atherogenesis by inhibiting disturbed flow-mediated endothelial activation via protein tyrosine phosphatase PTPN14 and YAP.

Disturbed flow induces endothelial dysfunction and contributes to uneven distribution of atherosclerotic plaque. Emerging evidence suggests that harmine, a natural constituent of extracts of Peganum harmala, has potent beneficial activities. Here, we investigated if harmine has an atheroprotective role under disturbed flow and the underlying mechanism. Mice of ApoE-/- , LDLR-/- , and endothelial cell (EC)-specific overexpression of yes-associated protein (YAP) in ApoE-/- background were fed with a Western diet and given harmine for 4 weeks. Atherosclerotic lesion size, cellular composition, and expression of inflammatory genes in the aortic roots were assessed. HUVECs were treated with oscillatory shear stress (OSS) and harmine and also used for proteomic analysis. Harmine retarded atherogenesis in both ApoE-/- and LDLR-/- mice by inhibiting the endothelial inflammatory response. Mechanistically, harmine blocked OSS-induced YAP nuclear translocation and EC activation by reducing phosphorylation of YAP at Y357. Overexpression of endothelial YAP blunted the beneficial effects of harmine in mice. Proteomic study revealed that protein tyrosine phosphatase non-receptor type 14 (PTPN14) could bind to YAP. Moreover, harmine increased PTPN14 expression by stabilizing its protein level and inhibiting its degradation in proteasomes. PTPN14 knockdown blocked the effects of harmine on YAPY357 and EC activation. Finally, overexpression of PTPN14 mimicked the effects of harmine and ameliorated atherosclerosis, and knockdown of PTPN14 blunted the atheroprotective effects of harmine and accelerated atherosclerosis, in a partial ligation mouse model. Harmine alleviated OSS-induced EC activation via a PTPN14/YAPY357 pathway and had a potent atheroprotective role.

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

ObjectivesAlterations 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...

Deletion of Alzheimer's disease-associated CD33 results in an inflammatory human microglia phenotype.

Genome-wide association studies demonstrated that polymorphisms in the CD33/sialic acid-binding immunoglobulin-like lectin 3 gene are associated with late-onset Alzheimer's disease (AD). CD33 is expressed on myeloid immune cells and mediates inhibitory signaling through protein tyrosine phosphatases, but the exact function of CD33 in microglia is still unknown. Here, we analyzed CD33 knockout human THP1 macrophages and human induced pluripotent stem cell-derived microglia for immunoreceptor tyrosine-based activation motif pathway activation, cytokine transcription, phagocytosis, and phagocytosis-associated oxidative burst. Transcriptome analysis of the macrophage lines showed that knockout of CD33 as well as knockdown of the CD33 signaling-associated protein tyrosine phosphatase, nonreceptor type 6 (PTPN6) led to constitutive activation of inflammation-related pathways. Moreover, deletion of CD33 or expression of Exon 2-deleted CD33 (CD33ΔE2 /CD33m) led to increased phosphorylation of the kinases spleen tyrosine kinase (SYK) and extracellular signal-regulated kinase 1 and 2 (ERK1 and 2). Transcript analysis by quantitative real-time polymerase chain reaction confirmed increased levels of interleukin (IL) 1B, IL8, and IL10 after knockout of CD33 in macrophages and microglia. In addition, upregulation of the gene transcripts of the AD-associated phosphatase INPP5D was observed after knockout of CD33. Functional analysis of macrophages and microglia showed that phagocytosis of aggregated amyloid-β1-42 and bacterial particles were increased after knockout of CD33 or CD33ΔE2 expression and knockdown of PTPN6. Furthermore, the phagocytic oxidative burst during uptake of amyloid-β1-42 or bacterial particles was increased after CD33 knockout but not in CD33ΔE2 -expressing microglia. In summary, deletion of CD33 or expression of CD33ΔE2 in human macrophages and microglia resulted in putative beneficial phagocytosis of amyloid β1-42 , but potentially detrimental oxidative burst and inflammation, which was absent in CD33ΔE2 -expressing microglia.

Titanium Dioxide Presents a Different Profile in Dextran Sodium Sulphate-Induced Experimental Colitis in Mice Lacking the IBD Risk Gene Ptpn2 in Myeloid Cells.

Environmental and genetic factors have been demonstrated to contribute to the development of inflammatory bowel disease (IBD). Recent studies suggested that the food additive; titanium dioxide (TiO2) might play a causative role in the disease. Therefore, in the present study we aimed to explore the interaction between the food additive TiO2 and the well-characterized IBD risk gene protein tyrosine phosphatase non-receptor type 2 (Ptpn2) and their role in the development of intestinal inflammation. Dextran sodium sulphate (DSS)-induced acute colitis was performed in mice lacking the expression of Ptpn2 in myeloid cells (Ptpn2LysMCre) or their wild type littermates (Ptpn2fl/fl) and exposed to the microparticle TiO2. The impact of Ptpn2 on TiO2 signalling pathways and TiO2-induced IL-1β and IL-10 levels were studied using bone marrow-derived macrophages (BMDMs). Ptpn2LysMCre exposed to TiO2 exhibited more severe intestinal inflammation than their wild type counterparts. This effect was likely due to the impact of TiO2 on the differentiation of intestinal macrophages, suppressing the number of anti-inflammatory macrophages in Ptpn2 deficient mice. Moreover, we also found that TiO2 was able to induce the secretion of IL-1β via mitogen-activated proteins kinases (MAPKs) and to repress the expression of IL-10 in bone marrow-derived macrophages via MAPK-independent pathways. This is the first evidence of the cooperation between the genetic risk factor Ptpn2 and the environmental factor TiO2 in the regulation of intestinal inflammation. The results presented here suggest that the ingestion of certain industrial compounds should be taken into account, especially in individuals with increased genetic risk

Molecular basis of the interaction of the human tyrosine phosphatase PTPN3 with the hepatitis B virus core protein

Interactions between the hepatitis B virus core protein (HBc) and host cell proteins are poorly understood, although they may be essential for the propagation of the virus and its pathogenicity. HBc has a C-terminal PDZ (PSD-95, Dlg1, ZO-1)-binding motif (PBM) that is responsible for interactions with host PDZ domain-containing proteins. In this work, we focused on the human protein tyrosine phosphatase non-receptor type 3 (PTPN3) and its interaction with HBc. We solved the crystal structure of the PDZ domain of PTPN3 in complex with the PBM of HBc, revealing a network of interactions specific to class I PDZ domains despite the presence of a C-terminal cysteine in this atypical PBM. We further showed that PTPN3 binds the HBc protein within capsids or as a homodimer. We demonstrate that overexpression of PTPN3 significantly affects HBV infection in HepG2 NTCP cells. Finally, we performed proteomics studies on both sides by pull-down assays and screening of a human PDZ domain library. We identified a pool of human PBM-containing proteins that might interact with PTPN3 in cells and that could be in competition with the HBc PBM during infection, and we also identified potential cellular partners of HBc through PDZ-PBM interactions. This study opens up many avenues of future investigations into the pathophysiology of HBV.

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.

Correlation Analysis of Th17/Treg Cells and PTPN22 Gene Polymorphism in HT Patients with Different Iodine Nutritional Status

Background: The incidence of Hashimoto's thyroiditis (HT) may be related to environmental, immune, and genetic factors. Different iodine nutritional status can affect thyroid function. Th17/Treg cell imbalance is involved in the pathogenesis of autoimmune thyroid diseases, and PTPN22 is an important susceptibility gene related to autoimmune diseases. To study the relationship between Th17/Treg cells and cytokines and PTPN22 gene polymorphism in HT patients with different iodine nutritional status is to determine the role of immune and genetic factors in the pathogenesis of HT. Objective: To investigate the correlation between Th17/Treg cells and factors protein tyrosine phosphatase non-receptor type 22 (PTPN22) gene-1123G>C site (rs2488457) polymorphism in peripheral blood mononuclear cells (PBMC) of Hashimoto's thyroiditis (HT) patients with different iodine nutritional status. Methods: 100 HT patients and 60 healthy subjects in Cang Zhou are selected from 2019 to 2020. Flow cytometry and real-time fluorescent quantitative PCR are used to detecting the proportion and ratio of Th17 and Treg cells in PBMC, as well as retinoic acid-related orphan receptor-γt (ROR-γt) and fork head/winged spiral transcription factor 3 (Foxp3) mRNA expression levels. Single allele-specific primer-polymerase chain reaction (SASP-PCR) technology is taken to sequence PTPN gene promoter-1123SNP leading to calculate the gene Type and allele frequency. Results: Th17 and Treg cells and factors in the PBMC of HT patients have a certain correlation with the-1123G>C locus of PTPN22 gene (rs2488457). The distribution frequency of rs2488457 genotype is consistent with Hardy Weinberg's law of genetic equilibrium. The distribution frequencies of various genotypes and alleles are significantly different between HT patients and the control group (P<0.05). PTPN22 gene-1123G>C site C/G, C/C genotype can increase of the risk of HT, while G/G genotype can decrease of the risk of HT. In Conclusion: The polymorphism of rs2488457 is related to susceptibility to HT. Carrying C/G and C/C genotypes may be risk factors for HT, and G/G genotypes may be protective factors for HT, but it has nothing to do with iodine nutrition status.

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.

Small‐Molecule Inhibitors of Shp2 Phosphatase as Potential Chemotherapeutic Agents for Glioblastoma: A Minireview

Glioblastoma multiforme (GBM) is a dreadful cancer characterised by poor prognosis, low survival rate and difficult clinical correlations. Several signalling pathways and molecular mediators are known to precipitate GBM, and small-molecular targets of these mediators have become a favoured thrust area for researchers to develop potent anti-GBM drugs. Shp2, an important phosphatase of the nonreceptor type protein tyrosine phosphatase (PTPN) subfamily is responsible for master regulation of several such signalling pathways in normal and glioma cells. Thus, inhibition of Shp2 is a logical strategy for the design and development of anti-neoplastic drugs against GBM. Though tapping the full potential of Shp2 binding sites has been challenging, nevertheless, many synthetic and natural scaffolds have been documented as possessing potent and selective anti-Shp2 activities in biochemical and cellular assays, through either active-site or allosteric binding. Most of these scaffolds share a few common pharmacophoric features, a thorough study of which is useful in paving the way for the design and development of improved Shp2 inhibitors. This minireview summarizes the current scenario of potent small-molecule Shp2 inhibitors and emphasizes the anti-GBM potential of some important scaffolds that have shown promising GBM-specific activity in in vitro and in vivo models, thus proving their efficacy in GBM therapy. This review could guide researchers to design new and improved anti-Shp2 pharmacophores and develop them as anti-GBM agents by employing GBM-centric drug-discovery protocols.

PTPN6 promotes chemosensitivity of colorectal cancer cells via inhibiting the SP1/MAPK signalling pathway.

The abnormal expression of protein tyrosine phosphatase nonreceptor type 6 (PTPN6) has been proved to be associated with the progression of colorectal cancer. However, its role in chemosensitivity and related molecular mechanism have not been clarified. It has been reported that PTPN6 was down-regulated in colorectal cancer cells compared with the normal colorectal cells. To evaluate the effects of PTPN6 on the proliferation and survival of colorectal cancer cells, PTPN6 was overexpressed in colorectal cancer cells in the present study. We found that cell proliferation and viability were both decreased after overexpression of PTPN6. The IC50 of 5-Fu against colorectal cells was also declined in PTPN6 transfected cells. And further, we verified that PTPN6 could down-regulate the expression of P-gp and MRP-1. Moreover, SP1 was the target protein of PTPN6 predicated by ChIPBase software and confirmed through Co-immunoprecipitation assay and it was negatively regulated by PTPN6. To further verify the effect of SP1 on chemoresistance, SP1 was overexpressed. SP1 overexpression enhanced the drug-resistance to 5-Fu and abrogated the effects of PTPN6 upregulation on 5-Fu resistance. All the above changes were associated with the down-regulation of proteins related to MAPK signalling pathway, such as phosphorylation of extracellular regulated protein kinases (ERK) and p38. In summary, PTPN6 promoted chemosensitivity of colorectal cancer cells by targeting SP1 and inhibiting the activation of MAPK signalling pathway. SIGNIFICANCE OF THE STUDY: It has been demonstrated that the abnormal expression of PTPN6 was related to the progression of colorectal cancer. However, the chemosensitivity of PTPN6 and its molecular mechanisms were still unclear. Here, we identified that PTPN6 was down-regulated in colorectal cancer cells. Moreover, PTPN6 overexpression not only reduced cell proliferation and viability, but decreased the resistance of colorectal cells to 5-Fu. In our research, we found that the SP1 was the target protein of PTPN6 and it was negatively regulated by PTPN6. In addition, SP1 could increase the resistance of colorectal cells to 5-Fu. Molecular mechanism studies have shown that PTPN6 promoted the chemosensitivity of colorectal cancer cells by inhibiting the activation of MAPK signalling pathway.

Tumour targetable and microenvironment-responsive nanoparticles simultaneously disrupt the PD-1/PD-L1 pathway and MAPK/ERK/JNK pathway for efficient treatment of colorectal cancer

The present study was aimed to develop a novel combination therapeutic strategy of gene therapy and immunotherapy for efficiently treatment of colorectal cancer (CRC). To achieve that goal, the polyethylene glycol-modified poly (2-(N,N-dimethylamino) ethyl methacrylate) (PEG-PDMAEMA)-based nanoparticles loaded with protein tyrosine phosphatase non-receptor type 6 (PTPN6) (NP-PTPN6) was developed first followed by conjugation with anti-PD-L1 monoclonal antibodies (aPD-L1) atezolizumab (aPD-L1NP-PTPN6). Importantly, the aPD-L1 was conjugated on the surface of NP-PTPN6 by the matrix metalloproteinases (MMPs)-cleavable linkage PLGLAG. Therefore, the aPD-L1 would be completely released once the aPD-L1NP-PTPN6 was entrapped into tumour tissues as demonstrated by the release assay. Tumour targeting assay demonstrated the aPD-L1NP-PTPN6 have high affinity to CRC cells and resulted in excellent tumour targeting drug delivery efficacy. Additionally, anti-tumour effect evaluation revealed that the aPD-L1NP-PTPN6 has greater ability to inhibit the growth, invasion and migration of CRC cells and finally led to longer survival time of tumour-bearing mice than other treatments. Further mechanisms studies demonstrated that treatment of CRC cells with aPD-L1NP-PTPN6 contributed to significant suppression of the MAPK/ERK signalling pathway. Besides, it was further demonstrated that treating CRC with aPD-L1NP-PTPN6 resulted in up-regulation of NK cells and T cells percentage within the tumour tissues.

Multi-color Molecular Visualization of Signaling Proteins Reveals How C-Terminal Src Kinase Nanoclusters Regulate T Cell Receptor Activation

Elucidating the mechanisms that controlled Tcell activation requires visualization of the spatial organization of multiple proteins on the submicron scale. Here, we use stoichiometrically accurate, multiplexed, single-molecule super-resolution microscopy (DNA-PAINT) to image the nanoscale spatial architecture of the primary inhibitor of the Tcell signaling pathway, Csk, and two binding partners implicated in its membrane association, PAG and TRAF3. Combined with a newly developed co-clustering analysis framework, we find that Csk forms nanoscale clusters proximal to the plasma membrane that are lost post-stimulation and are re-recruited at later time points. Unexpectedly, these clusters do not co-localize with PAG at the membrane but instead provide a ready pool of monomers to downregulate signaling. By generating CRISPR-Cas9 knockout Tcells, our data also identify that a major risk factor for autoimmune diseases, the protein tyrosine phosphatase non-receptor type 22 (PTPN22) locus, is essential for Csk nanocluster re-recruitment and for maintenance of the synaptic PAG population.

DNA hypermethylation of PTPN22 gene promoter in children and adolescents with Hashimoto thyroiditis

Protein tyrosine phosphatase non-receptor type 22 (PTPN22) is an inhibitor of T-cell activation, regulating intracellular signal transduction and thereby being implicated in the pathogenesis of autoimmune thyroid disease (AITD). The exact molecular mechanisms have not been fully elucidated. The aim of the present study was to quantitate DNA methylation within the PTPN22 gene promoter in children and adolescents with AITD and healthy controls. 60 Patients with Hashimoto thyroiditis (HT), 25 patients with HT and type 1 diabetes (HT + T1D), 9 patients with Graves' disease (GD) and 55 healthy controls without any individual or family history of autoimmune disease were enrolled. Whole blood DNA extraction, DNA modification using sodium bisulfate and quantification of DNA methylation in the PTPN22 gene promoter, based on melting curve analysis of the selected DNA fragment using a Real-Time PCR assay, were implemented. DNA methylation in the PTPN22 gene promoter was found to be significantly higher in HT patients (39.9 ± 3.1%) in comparison with other study groups (20.3 ± 2.4% for HT + T1D, 32.6 ± 7.8% for GD, 27.1 ± 2.4% for controls, p < 0.001). PTPN22 gene promoter DNA methylation was also associated marginally with thyroid autoimmunity in general (p = 0.059), as well as considerably with thyroid volume (p = 0.004) and the presence of goiter (p = 0.001) but not thyroid function tests. This study demonstrates for the first time that a relationship between autoimmune thyroiditis and PTPN22 gene promoter DNA methylation state is present, thus proposing another possible etiological association between thyroiditis and abnormalities of PTPN22 function. Further expression studies are required to confirm these findings.

SOX2 promotes resistance of melanoma with PD-L1 high expression to T-cell-mediated cytotoxicity that can be reversed by SAHA

BackgroundImmune checkpoint inhibitors (ICIs) induce better tumor regression in melanoma with programmed cell death 1 ligand 1 (PD-L1) high expression, but there has been an upsurge of failed responses. In...