ABSTRACT
Nuclear matrix protein 22 (NMP22) is one of the most important tumor markers of bladder cancer and is significantly elevated in the urine of bladder cancer patients. Therefore, in this work, a highly sensitive ratiometric electrochemical immunosensor was constructed to detect NMP22 based on ZIF-8@MWCNTs@Chit@Fc@AuNPs composites. ZIF-8 had a large surface area and good adsorption ability. Multi-Walled Carbon Nanotubes (MWCNTs) can optimize the electrical conductivity of ZIF-8, so that the electrode surface of ferrocene (Fc) obtains a stable and strong electrochemical signal. In addition, AuPt-MB provided another strong detection signal methylene blue (MB) while immobilizing the secondary antibody (Ab2) through Au-N and Pt-N bonds. A ratiometric electrochemical sensor was formed based on ZIF-8@MWCNTs@Chit@Fc@AuNPs and AuPt-MB, which showed a great linear connection between IMB/IFc and the logarithmic concentration of NMP22 with a detection limit of 3.33 fg mL-1 (S/N = 3) under optimized specifications in the concentration interval of 0.01 pg mL-1 to 1000 ng mL-1. In addition, the ratiometric immunosensor showed good selectivity and stability.
Subject(s)
Biosensing Techniques , Electrochemical Techniques , Gold , Metal Nanoparticles , Nanotubes, Carbon , Nuclear Proteins , Nanotubes, Carbon/chemistry , Gold/chemistry , Electrochemical Techniques/methods , Humans , Metal Nanoparticles/chemistry , Immunoassay/methods , Biosensing Techniques/methods , Nuclear Proteins/urine , Nuclear Proteins/immunology , Nuclear Proteins/analysis , Limit of Detection , Platinum/chemistry , Zeolites/chemistry , Methylene Blue/chemistry , Ferrous Compounds/chemistry , Antibodies, Immobilized/chemistry , Antibodies, Immobilized/immunology , Metallocenes/chemistry , Biomarkers, Tumor/urine , Biomarkers, Tumor/immunologyABSTRACT
Single-stranded DNA containing CGT/A motifs binds to the helicase domain of Schlafen 11 (SLFN11) to initiate cell death and cytokine production via SLFN11 ribonuclease activity (see related Research Article by Zhang et al.).
Subject(s)
DNA, Single-Stranded , Immunity, Innate , Animals , Humans , DNA, Single-Stranded/immunology , DNA, Single-Stranded/metabolism , Immunity, Innate/immunology , Nuclear Proteins/immunology , Nuclear Proteins/metabolism , Ribonucleases/immunology , Ribonucleases/metabolismABSTRACT
Nucleic acids are major structures detected by the innate immune system. Although intracellular single-stranded DNA (ssDNA) accumulates during pathogen infection or disease, it remains unclear whether and how intracellular ssDNA stimulates the innate immune system. Here, we report that intracellular ssDNA triggers cytokine expression and cell death in a CGT motif-dependent manner. We identified Schlafen 11 (SLFN11) as an ssDNA-activated RNase, which is essential for the innate immune responses induced by intracellular ssDNA and adeno-associated virus infection. We found that SLFN11 directly binds ssDNA containing CGT motifs through its carboxyl-terminal domain, translocates to the cytoplasm upon ssDNA recognition, and triggers innate immune responses through its amino-terminal ribonuclease activity that cleaves transfer RNA (tRNA). Mice deficient in Slfn9, a mouse homolog of SLFN11, exhibited resistance to CGT ssDNA-induced inflammation, acute hepatitis, and septic shock. This study identifies CGT ssDNA and SLFN11/9 as a class of immunostimulatory nucleic acids and pattern recognition receptors, respectively, and conceptually couples DNA immune sensing to controlled RNase activation and tRNA cleavage.
Subject(s)
DNA, Single-Stranded , Immunity, Innate , Mice, Inbred C57BL , Animals , Female , Humans , Male , Mice , DNA, Single-Stranded/immunology , HEK293 Cells , Immunity, Innate/immunology , Mice, Knockout , Nuclear Proteins/immunology , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Ribonucleases/immunology , Ribonucleases/metabolismABSTRACT
A 23-year-old man was admitted to our hospital with a one-year history of muscle weakness and atrophy. He had noticed contractures of the fingers of both hands from the age of 18. Examination revealed a skin rash including heliotrope rash and Gottron's sign, joint contractures in the extremities, dysphagia, extensive muscle weakness and marked muscle atrophy. The serum creatine kinase level was 272â |IU/l and muscle biopsy showed typical perifascicular atrophy but little lymphocyte invasion. There was no interstitial pneumonia or malignancy, but muscle tendons showed elevated CT values suggesting calcification or fibrosis. Anti-nuclear matrix protein 2 (NXP-2) antibody-positive dermatomyositis was diagnosed on the basis of the serum antibody level. Methylprednisolone pulse therapy ameliorated the skin rash and bulbar palsy, but muscle weakness, atrophy and joint contractures were resistant to the treatment. There have been no previous reports of young adults with anti-NXP-2 antibody-positive dermatomyositis in whom joint contracture became evident as early as 4 years beforehand, which is a important feature for differential diagnosis of dermatomyositis.
Subject(s)
Autoantibodies , Biomarkers , Contracture , Dermatomyositis , Pulse Therapy, Drug , Humans , Male , Young Adult , Adenosine Triphosphatases , Autoantibodies/blood , Biomarkers/blood , Contracture/etiology , Contracture/diagnosis , Dermatomyositis/complications , Dermatomyositis/immunology , Dermatomyositis/diagnosis , Dermatomyositis/drug therapy , Diagnosis, Differential , DNA-Binding Proteins , Methylprednisolone/administration & dosage , Nuclear Proteins/immunology , RNA-Binding Proteins/immunology , Transcription FactorsABSTRACT
Maternal infection during pregnancy is an important cause of autism spectrum disorder (ASD) in offspring, and inflammatory infiltration caused by maternal immune activation (MIA) can cause neurodevelopmental disorders in the fetus. Medicine food homologous (MFH) refers to a traditional Chinese medicine (TCM) concept, which effectively combines food functions and medicinal effects. However, no previous study has screened, predicted, and validated the potential targets of MFH herbs for treating ASD. Therefore, in this study, we used comprehensive bioinformatics methods to screen and analyze MFH herbs and drug targets on a large scale, and identified resveratrol and Thoc5 as the best small molecular ingredient and drug target, respectively, for the treatment of MIA-induced ASD. Additionally, the results of in vitro experiments revealed that resveratrol increased the expression of Thoc5 and effectively inhibited lipopolysaccharide-induced inflammatory factor production by BV2 cells. Moreover, in vivo, resveratrol increased the expression of Thoc5 and effectively inhibited placental and fetal brain inflammation in MIA pregnancy mice, and improved ASD-like behaviors in offspring.
Subject(s)
Autism Spectrum Disorder , Nuclear Proteins , Prenatal Exposure Delayed Effects , Resveratrol , Animals , Female , Male , Mice , Pregnancy , Autism Spectrum Disorder/immunology , Autistic Disorder/chemically induced , Autistic Disorder/immunology , Behavior, Animal/drug effects , Disease Models, Animal , Lipopolysaccharides/toxicity , Mice, Inbred C57BL , Resveratrol/pharmacology , Nuclear Proteins/drug effects , Nuclear Proteins/immunology , Nuclear Proteins/metabolismABSTRACT
DNA replication and sister chromatid cohesion 1 (DSCC1) exerts various functions including sister chromatid cohesion. DSCC1 overexpression plays an important role in cancer development, such as in colorectal, breast, and hepatocellular cancers. The specific role of DSCC1 in tumor progression remains largely unknown, necessitating a pan-cancer investigation to understand the potential function of DSCC1 in various cancers. In this study, we obtained data on physiological conditions, transcriptional expression, survival prognosis, genomic alteration, genomic instability, enriched pathways, immune infiltration, and immunotherapy from The Cancer Genome Atlas, The Genotype-Tissue Expression, cBioPortal, and other publicly available databases to systematically characterize the oncogenic and immunological roles of DSCC1 in 33 different cancers. We found that DSCC1 expression was upregulated at both mRNA and protein levels in various cancers. Additionally, DSCC1 expression was associated with higher tumor stage and grade in specific cancers. DSCC1 was a potential pan-cancer prognostic biomarker for its close association with patient prognosis and a diagnostic biomarker for its high predictive value in distinguishing tumor tissues from normal tissues. DSCC1 was universally amplified across different cancers and tightly associated with genomic instability. Moreover, DSCC1 had a close relationship with tumor immune cell infiltration; thus, it could be used as a potential biomarker for predicting the response and survival of patients with cancer who receive immune checkpoint blockade treatment. To sum up, our study revealed that DSCC1 is a promising target for tumor therapy.
Subject(s)
Biomarkers, Tumor , Gene Expression Regulation, Neoplastic , Genomic Instability , Neoplasms , Nuclear Proteins , Humans , Biomarkers, Tumor/genetics , Immunotherapy , Neoplasms/genetics , Neoplasms/immunology , Neoplasms/diagnosis , Prognosis , Nuclear Proteins/genetics , Nuclear Proteins/immunologyABSTRACT
PYHIN proteins are only found in mammals and play key roles in the defense against bacterial and viral pathogens. The corresponding gene locus shows variable deletion and expansion ranging from 0 genes in bats, over 1 in cows, and 4 in humans to a maximum of 13 in mice. While initially thought to act as cytosolic immune sensors that recognize foreign DNA, increasing evidence suggests that PYHIN proteins also inhibit viral pathogens by more direct mechanisms. Here, we examined the ability of all 13 murine PYHIN proteins to inhibit HIV-1 and murine leukemia virus (MLV). We show that overexpression of p203, p204, p205, p208, p209, p210, p211, and p212 strongly inhibits production of infectious HIV-1; p202, p207, and p213 had no significant effects, while p206 and p214 showed intermediate phenotypes. The inhibitory effects on infectious HIV-1 production correlated significantly with the suppression of reporter gene expression by a proviral Moloney MLV-eGFP construct and HIV-1 and Friend MLV LTR luciferase reporter constructs. Altogether, our data show that the antiretroviral activity of PYHIN proteins is conserved between men and mice and further support the key role of nuclear PYHIN proteins in innate antiviral immunity.
Subject(s)
HIV-1 , Leukemia Virus, Murine , Phosphoproteins , Animals , Mice , Humans , HIV-1/immunology , HIV-1/genetics , Leukemia Virus, Murine/genetics , Leukemia Virus, Murine/immunology , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Nuclear Proteins/immunology , Virus Replication , Cell Line , Retroviridae Infections/immunology , Retroviridae Infections/virologyABSTRACT
BACKGROUND: Colorectal cancer (CRC) is the third most common of cancer-related deaths. Nucleolar protein 14 (NOP14) is known to play different roles in diverse types of cancers. However, little is known about its roles in CRC. Here, we assessed the prognostic value and functions of NOP14 in CRC using the data from The Cancer Genome Atlas (TCGA) and validated them based on the data from Gene Expression Omnibus (GEO). METHODS: NOP14 mRNA and protein data in CRC was obtained from the TCGA, GEO, human protein atlas (HPA), and UALCAN databases. Survival and Cox regression analysis was performed to assess the prognostic value of NOP14 in CRC patients. Next, to evaluate the potential functions of NOP14, a protein-protein interaction (PPI) network was constructed and gene set enrichment analysis (GSEA) of differential expression genes (DEGs) associated with dysregulated NOP14 was performed. Finally, to investigate the immune response associated with NOP14 expression in CRC, we analyzed the correlations between immune cells infiltration and NOP14 expression level. Additionally, the correlations between immune molecule expression levels with NOP14 expression level were analyzed. RESULTS: High NOP14 mRNA expression was observed in CRC tissues based on the data from TCGA and GEO datasets. Similarly, high NOP14 protein levels were found in CRC tissues according to the immunohistochemical images from HPA. Interestingly, high NOP14 expression level was associated with an improved prognosis in CRC patients. Univariate and multivariate Cox regression analysis indicated that high NOP14 expression level was an independent protective factor for CRC patients. With the support of PPI network analysis, we found several risk genes interacted with NOP14. GSEA revealed that high NOP14 expression inhibited several signal pathways involved in tumor formation and development. Additionally, high NOP14 expression was positively associated with most kinds of immune cell infiltrations and the expression levels of some molecules related to immune activation. CONCLUSION: Altogether, these results indicated that high NOP14 expression leads to improved prognosis in CRC patients by inhibiting the signaling pathways involved in tumor growth and promoting the immune responses.
Subject(s)
Colorectal Neoplasms , Nuclear Proteins , Biomarkers, Tumor/biosynthesis , Biomarkers, Tumor/genetics , Biomarkers, Tumor/immunology , Colorectal Neoplasms/genetics , Colorectal Neoplasms/immunology , Colorectal Neoplasms/pathology , Humans , Nuclear Proteins/genetics , Nuclear Proteins/immunology , Prognosis , RNA, Messenger/genetics , RNA, Messenger/immunologyABSTRACT
BACKGROUND: Thoracic SMARCA4-deficient undifferentiated tumors (SMARCA4-UT) are aggressive neoplasms. Data linking BAF alterations with tumor microenvironment (TME) and efficacy of immune checkpoint inhibitors (ICI) are contradictory. The TME of SMARCA4-UT and their response to ICI are unknown. MATERIALS AND METHODS: Patients diagnosed with SMARCA4-UT in our institution were included. Immunostainings for tertiary lymphoid structures (TLS), immune cell markers, and checkpoints were assessed. Validation was performed using an independent transcriptome dataset including SMARCA4-UT, non-small cell lung cancers (NSCLC) with/without SMARCA4 mutations, and unclassified thoracic sarcomas (UTS). CXCL9 and PD-L1 expressions were assessed in NSCLC and thoracic fibroblast cell lines, with/without SMARCA4 knockdown, treated with/without interferon gamma. RESULTS: Nine patients were identified. All samples but one showed no TLS, consistent with an immune desert TME phenotype. Four patients received ICI as part of their treatment, but the only one who responded, had a tumor with a TLS and immune-rich TME. Unsupervised clustering of the validation cohort using immune cell scores identified 2 clusters associated with cell ontogeny and immunity (cluster 1 enriched for NSCLC independently of SMARCA4 status (n = 9/10; P = .001); cluster 2 enriched for SMARCA4-UT (n = 11/12; P = .005) and UTS (n = 5/5; P = .0005). SMARCA4 loss-of-function experiments revealed interferon-induced upregulation of CXCL9 and PD-L1 expression in the NSCLC cell line with no effect on the thoracic fibroblast cell line. CONCLUSION: SMARCA4-UT mainly have an immune desert TME with limited efficacy to ICI. TME of SMARCA4-driven tumors varies according to the cell of origin questioning the interplay between BAF alterations, cell ontogeny and immunity.
Subject(s)
Carcinoma, Non-Small-Cell Lung , DNA Helicases , Immune Checkpoint Inhibitors , Lung Neoplasms , Nuclear Proteins , Sarcoma , Soft Tissue Neoplasms , Thoracic Neoplasms , B7-H1 Antigen/antagonists & inhibitors , B7-H1 Antigen/immunology , Biomarkers, Tumor/immunology , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/immunology , Carcinoma, Non-Small-Cell Lung/pathology , DNA Helicases/deficiency , DNA Helicases/immunology , Humans , Immune Checkpoint Inhibitors/pharmacology , Lung Neoplasms/drug therapy , Lung Neoplasms/immunology , Lung Neoplasms/pathology , Nuclear Proteins/deficiency , Nuclear Proteins/immunology , Sarcoma/drug therapy , Sarcoma/immunology , Sarcoma/pathology , Soft Tissue Neoplasms/drug therapy , Soft Tissue Neoplasms/immunology , Soft Tissue Neoplasms/pathology , Thoracic Neoplasms/drug therapy , Thoracic Neoplasms/immunology , Thoracic Neoplasms/pathology , Transcription Factors/immunology , Tumor Microenvironment/immunologyABSTRACT
N6-methyladenosine (m6A) has been reported as an important mechanism of post-transcriptional regulation. Programmed death ligand 1 (PD-L1) is a primary immune inhibitory molecule expressed on tumor cells that promotes immune evasion. In addition, seven in absentia homolog 2 (Siah2), a RING E3 ubiquitin ligase, has been involved in tumorigenesis and cancer progression. However, the role of m6A-METTL14-Siah2-PD-L1 axis in immunotherapy remains to be elucidated. In this study, we showed that METTL14, a component of the m6A methyltransferase complex, induced Siah2 expression in cholangiocarcinoma (CCA). METTL14 was shown to enrich m6A modifications in the 3'UTR region of the Siah2 mRNA, thereby promoting its degradation in an YTHDF2-dependent manner. Furthermore, co-immunoprecipitation experiments demonstrated that Siah2 interacted with PD-L1 by promoting its K63-linked ubiquitination. We also observed that in vitro and in vivo Siah2 knockdown inhibited T cells expansion and cytotoxicity by sustaining tumor cell PD-L1 expression. The METTL14-Siah2-PD-L1-regulating axis was further confirmed in human CCA specimens. Analysis of specimens from patients receiving anti-PD1 immunotherapy suggested that tumors with low Siah2 levels were more sensitive to anti-PD1 immunotherapy. Taken together, our results evidenced a new regulatory mechanism of Siah2 by METTL14-induced mRNA epigenetic modification and the potential role of Siah2 in cancer immunotherapy.
Subject(s)
B7-H1 Antigen/immunology , Cholangiocarcinoma/immunology , Nuclear Proteins/immunology , T-Lymphocytes/immunology , Ubiquitin-Protein Ligases/immunology , Adenosine/analogs & derivatives , Adenosine/immunology , Cell Line , Cholangiocarcinoma/therapy , Humans , Immunotherapy , Methyltransferases/immunology , RNA, Messenger/immunologyABSTRACT
Human cytomegalovirus (HCMV) is an important human pathogen and a paradigm of viral immune evasion, targeting intrinsic, innate, and adaptive immunity. We have employed two orthogonal multiplexed tandem mass tag-based proteomic screens to identify host proteins down-regulated by viral factors expressed during the latest phases of viral infection. This approach revealed that the HIV-1 restriction factor Schlafen-11 (SLFN11) was degraded by the poorly characterized, late-expressed HCMV protein RL1, via recruitment of the Cullin4-RING E3 Ubiquitin Ligase (CRL4) complex. SLFN11 potently restricted HCMV infection, inhibiting the formation and spread of viral plaques. Overall, we show that a restriction factor previously thought only to inhibit RNA viruses additionally restricts HCMV. We define the mechanism of viral antagonism and also describe an important resource for revealing additional molecules of importance in antiviral innate immunity and viral immune evasion.
Subject(s)
Cytomegalovirus Infections/immunology , Cytomegalovirus/immunology , Immune Evasion , Nuclear Proteins/immunology , Proteolysis , Viral Envelope Proteins/immunology , Cytomegalovirus/genetics , Cytomegalovirus Infections/genetics , Humans , Nuclear Proteins/genetics , Ubiquitin-Protein Ligase Complexes/genetics , Ubiquitin-Protein Ligase Complexes/immunology , Viral Envelope Proteins/geneticsABSTRACT
Virus infection triggers intricate signal cascade reactions to activate the host innate immunity, which leads to the production of type I interferon (IFN-I). Herpes simplex virus 1 (HSV-1), a human-restricted pathogen, is capable of encoding over 80 viral proteins, and several of them are involved in immune evasion to resist the host antiviral response through the IFN-I signaling pathway. Here, we determined that HSV-1 UL31, which is associated with nuclear matrix and is essential for the formation of viral nuclear egress complex, could inhibit retinoic acid-inducible gene I (RIG-I)-like receptor pathway-mediated interferon beta (IFN-ß)-luciferase (Luc) and (PRDIII-I)4-Luc (an expression plasmid of IFN-ß positive regulatory elements III and I) promoter activation, as well as the mRNA transcription of IFN-ß and downstream interferon-stimulated genes (ISGs), such as ISG15, ISG54, ISG56, etc., to promote viral infection. UL31 was shown to restrain IFN-ß activation at the interferon regulatory factor 3 (IRF3)/IRF7 level. Mechanically, UL31 was demonstrated to interact with TANK binding kinase 1 (TBK1), inducible IκB kinase (IKKi), and IRF3 to impede the formation of the IKKi-IRF3 complex but not the formation of the IRF7-related complex. UL31 could constrain the dimerization and nuclear translocation of IRF3. Although UL31 was associated with the CREB binding protein (CBP)/p300 coactivators, it could not efficiently hamper the formation of the CBP/p300-IRF3 complex. In addition, UL31 could facilitate the degradation of IKKi and IRF3 by mediating their K48-linked polyubiquitination. Taken together, these results illustrated that UL31 was able to suppress IFN-ß activity by inhibiting the activation of IKKi and IRF3, which may contribute to the knowledge of a new immune evasion mechanism during HSV-1 infection. IMPORTANCE The innate immune system is the first line of host defense against the invasion of pathogens. Among its mechanisms, IFN-I is an essential cytokine in the antiviral response, which can help the host eliminate a virus. HSV-1 is a double-stranded DNA virus that can cause herpes and establish a lifelong latent infection, due to its possession of multiple mechanisms to escape host innate immunity. In this study, we illustrate for the first time that the HSV-1-encoded UL31 protein has a negative regulatory effect on IFN-ß production by blocking the dimerization and nuclear translocation of IRF3, as well as promoting the K48-linked polyubiquitination and degradation of both IKKi and IRF3. This study may be helpful for fully understanding the pathogenesis of HSV-1.
Subject(s)
Herpesvirus 1, Human/genetics , Herpesvirus 1, Human/immunology , Interferon-beta/genetics , Interferon-beta/immunology , Nuclear Proteins/genetics , Nuclear Proteins/immunology , Viral Proteins/genetics , Viral Proteins/immunology , Animals , Chlorocebus aethiops , Cytokines , DEAD Box Protein 58 , HEK293 Cells , HeLa Cells , Herpes Simplex , Host-Pathogen Interactions , Humans , Immune Evasion , Immunity, Innate , Interferon Regulatory Factor-3/genetics , Interferon Regulatory Factor-3/immunology , Interferon Regulatory Factor-3/metabolism , Interferon Regulatory Factor-7 , Interferon Type I , Interferon-beta/metabolism , Nuclear Proteins/metabolism , Protein Serine-Threonine Kinases , Receptors, Immunologic , Signal Transduction , Vero Cells , Viral Proteins/metabolismABSTRACT
The Bacillus Calmette-Guérin (BCG) vaccine is known to have protective effects not only against tuberculosis but also against other unrelated infectious diseases caused by different pathogens. Several epidemiological studies have also documented the beneficial influence of BCG vaccine in reducing both susceptibility to and severity of SARS-CoV-2 infection. The protective, non-specific effects of BCG vaccination would be related to an antigen-independent enhancement of the innate immunity, termed trained immunity. However, the knowledge that heat shock protein (HSP)65 is the main antigen of Mycobacterium bovis BCG prompted us to verify whether sequence similarity existed between HSP65 and SARS-CoV-2 spike (S) and nuclear (N) proteins that could support an antigen-driven immune protection of BCG vaccine. The results of the in silico investigation showed an extensive sequence similarity of HSP65 with both the viral proteins, especially SARS-CoV-2 S, that also involved the regions comprising immunodominant epitopes. The finding that the predicted B cell and CD4+ T cell epitopes of HSP65 shared strong similarity with the predicted B and T cell epitopes of both SARS-CoV-2 S and N would support the possibility of a cross-immune reaction of HSP65 of BCG with SARS-CoV-2.
Subject(s)
BCG Vaccine/immunology , COVID-19/immunology , Heat-Shock Proteins/immunology , Immunity, Innate/immunology , Mycobacterium bovis/virology , BCG Vaccine/pharmacology , COVID-19/prevention & control , Humans , Mycobacterium bovis/immunology , Nuclear Proteins/immunology , SARS-CoV-2/immunologySubject(s)
Autoantibodies/immunology , Carcinoma, Squamous Cell/immunology , Nuclear Proteins/immunology , RNA-Binding Proteins/immunology , Scleroderma, Systemic/immunology , Skin Neoplasms/immunology , Adult , Aged , Autoantigens/immunology , Female , Humans , Male , Middle Aged , Scleroderma, Systemic/complicationsABSTRACT
OBJECTIVE: Double-strand (ds) DNA-enveloped viruses can cause oral infection. Our aim is to investigate whether oral mucosal cells participate in immune response against cytosolic dsDNA invasion. METHODS: We examined the response to transfected herpes simplex virus (HSV) dsDNA via intracellular receptors in oral keratinocytes (RT7) and fibroblasts (GT1), and the effect of TNF-α on those responses. RESULTS: Transfected dsDNA increased CXCL10 expression via NF-κB activation in both cell types, while those responses were inhibited by knockdown of RIG-I, an RNA sensor. Although IFI16, a DNA sensor, was expressed in the nuclei of both types, its knockdown decreased transfected dsDNA-induced CXCL10 expression in GT1 but not RT7 cells. IFI16 in GT1 cells was translocated into cytoplasm from nuclei, which was attributed to immune response to cytosolic dsDNA. TNF-α enhanced transfected dsDNA-induced CXCL10, and knockdown of IFI16 decreased TNF-α and dsDNA-driven CXCL10 expression in both RT7 and GT1 cells. Finally, the combination of TNF-α and transfected dsDNA resulted in translocation of IFI16 from nuclei to cytoplasm in RT7 cells. CONCLUSION: RIG-I and IFI16 in oral mucosal cells may play important roles in host immune response against DNA viral infection, while TNF-α contributes to development of an antiviral system via those intracellular receptors.
Subject(s)
DNA, Viral/immunology , Fibroblasts , Keratinocytes , Simplexvirus/immunology , Antiviral Restriction Factors/immunology , Cell Line , Chemokine CXCL10/immunology , Cytoplasm , Fibroblasts/immunology , Humans , Immunity , Keratinocytes/immunology , Nuclear Proteins/immunology , Phosphoproteins/immunology , Receptors, Retinoic Acid/immunology , Tumor Necrosis Factor-alpha/immunologyABSTRACT
Interferon gamma-inducible protein 16 (IFI16) is a DNA sensor protein, which triggers interferon-beta (IFN-ß) production. However, the role of IFI16 in the innate immunity against hepatitis B virus (HBV) remains controversial. Peripheral blood mononuclear cells (PBMCs) and serum specimens were collected from 20 patients with chronic hepatitis B (CHB) receiving Peg-IFN-α2b therapy. IFI16 mRNA/protein of PBMCs and serum IFI16 at baseline and changes during Peg-IFN-α2b treatment were detected. The interaction between IFI16 and HBV DNA in the PBMCs was analyzed using chromatin immunoprecipitation assay. Leukemic T cell line CEM-C7 and HBV-replicating HepG2.2.15 cells were used to test the effects of interferon treatment and HBV replication on IFI16 expression. Compared with healthy controls, lower levels of IFI16 mRNA but more significant expression of IFI16 protein with heterogeneous degradation were detected in PBMCs of CHB patients. Early changes in IFI16 mRNA, but not IFNB mRNA of PBMCs or serum IFI16, were correlated to HBeAg seroconversion of Peg-IFN-α2b therapy. An interaction between IFI16 and HBV DNA was detected in the PBMCs. In the cultured HepG2.2.15 and CEM-C7 cells, interferons resulted in the translocalization of IFI16 from the cytoplasm to the nucleus and inhibited IFI16 degradation. IFI16 of PBMCs may play a role in sensing HBV infection, and early change in IFI16 mRNA of PBMCs is valuable to predict HBeAg seroconversion in Peg-IFN-α2b treatment. The influences on IFI16 degradation and subcellular location may present a molecular mechanism of antiviral activity of interferon.
Subject(s)
Antiviral Agents , Hepatitis B , Nuclear Proteins/immunology , Phosphoproteins/immunology , Antiviral Agents/therapeutic use , Cells, Cultured , Hepatitis B/drug therapy , Hepatitis B/immunology , Humans , Interferon alpha-2/therapeutic use , Interferon-alpha/therapeutic use , Leukocytes, Mononuclear , Polyethylene Glycols/therapeutic use , Recombinant Proteins/therapeutic useABSTRACT
BACKGROUND: Induction of CD8+ T cells that recognize immunogenic, mutated protein fragments in the context of major histocompatibility class I (MHC-I) is a pressing challenge for cancer vaccine development. METHODS: Using the commonly used murine renal adenocarcinoma RENCA cancer model, MHC-I restricted neoepitopes are predicted following next-generation sequencing. Candidate neoepitopes are screened in mice using a potent cancer vaccine adjuvant system that converts short peptides into immunogenic nanoparticles. An identified functional neoepitope vaccine is then tested in various therapeutic experimental tumor settings. RESULTS: Conversion of 20 short MHC-I restricted neoepitope candidates into immunogenic nanoparticles results in antitumor responses with multivalent vaccination. Only a single neoepitope candidate, Nesprin-2 L4492R (Nes2LR), induced functional responses but still did so when included within 20-plex or 60-plex particles. Immunization with the short Nes2LR neoepitope with the immunogenic particle-inducing vaccine adjuvant prevented tumor growth at doses multiple orders of magnitude less than with other vaccine adjuvants, which were ineffective. Nes2LR vaccination inhibited or eradicated disease in subcutaneous, experimental lung metastasis and orthotopic tumor models, synergizing with immune checkpoint blockade. CONCLUSION: These findings establish the feasibility of using short, MHC-I-restricted neoepitopes for straightforward immunization with multivalent or validated neoepitopes to induce cytotoxic CD8+ T cells. Furthermore, the Nes2LR neoepitope could be useful for preclinical studies involving renal cell carcinoma immunotherapy.
Subject(s)
Antigens, Neoplasm/immunology , CD8-Positive T-Lymphocytes/immunology , Cancer Vaccines/administration & dosage , Carcinoma, Renal Cell/prevention & control , Epitopes/immunology , Nerve Tissue Proteins/immunology , Nuclear Proteins/immunology , Peptide Fragments/pharmacology , Animals , Carcinoma, Renal Cell/immunology , Carcinoma, Renal Cell/pathology , Female , Histocompatibility Antigens Class I/immunology , Kidney Neoplasms/immunology , Kidney Neoplasms/pathology , Kidney Neoplasms/prevention & control , Lung Neoplasms/immunology , Lung Neoplasms/prevention & control , Lung Neoplasms/secondary , Mice , Mice, Inbred BALB C , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Peptide Fragments/immunology , T-Lymphocytes, Cytotoxic/immunologyABSTRACT
Breast cancer (BC) is a heterogeneous disease composed of multiple subtypes with different molecular characteristics and clinical outcomes. The metastatic process in BC depends on the transcription factors (TFs) related to epithelial-mesenchymal transition (EMT), including the master regulator Twist1. However, its role beyond EMT in BC subtypes remains unclear. Our study aimed to investigate the role of Twist1, beyond EMT, in the molecular subtypes of BC. In patients, we observed the overexpression of TWIST1 in the HER2+ group. The silencing of TWIST1 in HER2+ BC cells resulted in the upregulation of 138 genes and the downregulation of 174 genes compared to control cells in a microarray assay. In silico analysis revealed correlations between Twist1 and important biological processes such as the Th17-mediated immune response, suggesting that Twist1 could be relevant for IL-17 signaling in HER2+ BC. IL-17 signaling was then examined, and it was shown that TWIST1 knockdown caused the downregulation of leading members of IL-17 signaling pathway. Taken together, our findings suggest that Twist1 plays a role on IL-17 signaling in HER2+ BC.
Subject(s)
Breast Neoplasms/immunology , Gene Expression Regulation, Neoplastic/immunology , Interleukin-17/immunology , Nuclear Proteins/immunology , Receptor, ErbB-2/immunology , Signal Transduction/immunology , Twist-Related Protein 1/immunology , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Cell Line, Tumor , Female , Humans , Interleukin-17/genetics , Nuclear Proteins/genetics , Receptor, ErbB-2/genetics , Signal Transduction/genetics , Twist-Related Protein 1/geneticsABSTRACT
IFN-γ-inducible protein 16 (IFI16) recognizes viral DNAs from both nucleus-replicating viruses and cytoplasm-replicating viruses. Isoform 2 of IFI16 (IFI16-iso2) with nuclear localization sequence (NLS) has been studied extensively as a well-known DNA sensor. However, the characteristics and functions of other IFI16 isoforms are almost unknown. Here, we find that IFI16-iso1, with exactly the same length as IFI16-iso2, lacks the NLS and locates in the cytoplasm. To distinguish the functions of IFI16-iso1 and IFI16-iso2, we have developed novel nuclear viral DNA mimics that can be recognized by the nuclear DNA sensors, including IFI16-iso2 and hnRNPA2B1. The hexanucleotide motif 5'-AGTGTT-3' DNA form of the nuclear localization sequence (DNLS) effectively drives cytoplasmic viral DNA nuclear translocation. These nuclear viral DNA mimics potently induce IFN-ß and antiviral IFN-stimulated genes in human A549 cells, HEK293T cells, and mouse macrophages. The subcellular location difference of IFI16 isoforms determines their differential functions in recognizing viral DNA and activating type I IFN-dependent antiviral immunity. IFI16-iso1 preferentially colocalizes with cytoplasmic HSV60mer and cytoplasm-replicating vaccinia virus (VACV), whereas IFI16-iso2 mainly colocalizes with nuclear HSV60-DNLS and nucleus-replicating HSV-1. Compared with IFI16-iso2, IFI16-iso1 induces more transcription of IFN-ß and IFN-stimulated genes, as well as stronger antiviral immunity upon HSV60mer transfection or VACV infection. IFI16-iso2, with the ability of nuclear-cytoplasmic shuttling, clears both invaded HSV type 1 and VACV significantly. However, IFI16-iso2 induces more type I IFN-dependent antiviral immunity than IFI16-iso1 upon HSV60-DNLS transfection or HSV type 1 infection. Our study has developed potent agonists for nuclear DNA sensors and also has demonstrated that IFI16 isoforms with cytoplasmic and nuclear locations play differential roles in innate immunity against DNA viruses.
Subject(s)
Cell Nucleus/immunology , DNA Viruses/immunology , Nuclear Proteins/immunology , Phosphoproteins/immunology , Cells, Cultured , Humans , Protein Isoforms/immunologyABSTRACT
The inflammasome pathway is an important arm of the innate immune system that provides antiviral immunity against many viruses. The main pathways involved in virus infections include the NLRP3, IFI16, and AIM2 pathways. However, a succinct understanding of its role in HIV is not yet well elucidated. In this review, we showed that NLRP3 inflammasome activation plays a vital role in inhibiting HIV entry into target cells via the purinergic pathway; IFI16 detects intracellular HIV ssDNA, triggers interferon I and III production, and inhibits HIV transcription; and AIM2 binds to HIV dsDNA and triggers acute inflammation and pyroptosis. Remarkably, by understanding these mechanisms, new therapeutic strategies can be developed against the disease.