Identification of RP11-770J1.4 as immune-related lncRNA regulating the CTXN1-cGAS-STING axis in histologically lower-grade glioma.

Human gliomas are lethal brain cancers. Emerging evidence revealed the regulatory role of long noncoding RNAs (lncRNAs) in tumors. Here, we performed a comprehensive analysis of the expression profiles of RNAs in histologically lower-grade glioma (LGG). Enrichment analysis revealed that glioma is influenced by immune-related signatures. Survival analysis further established the close correlation between network features and glioma prognosis. Subsequent experiments showed lncRNA RP11-770J1.4 regulates CTXN1 expression through hsa-miR-124-3p. Correlation analysis identified lncRNA RP11-770J1.4 was immune related, specifically involved in the cytosolic DNA sensing pathway. Downregulated lncRNA RP11-770J1.4 resulted in increased spontaneous gene expression of the cGAS-STING pathway. Single-cell RNA sequencing analysis, along with investigations in a glioblastoma stem cell model and patient sample analysis, demonstrated the predominant localization of CTXN1 within tumor cores rather than peripheral regions. Immunohistochemistry staining established a negative correlation between CTXN1 expression and infiltration of CD8+ T cells. In vivo, Ctxn1 knockdown in GL261 cells led to decreased tumor burden and improved survival while increasing infiltration of CD8+ T cells. These findings unveil novel insights into the lncRNA RP11-770J1.4-CTXN1 as a potential immune regulatory axis, highlighting its therapeutic implications for histologically LGGs.

Progress in DNA-sensing pathways in Epstein-Barr virus infection / 中华微生物学和免疫学杂志

Epstein-Barr virus (EBV) is generally susceptible in human beings and multi-organ systems can be involved in EBV infection, such as blood, respiratory, urinary, digestive and nervous systems. EBV infection also plays an important role in the pathogenesis of related tumors, autoimmune diseases and other diseases, posing a great threat to human health. As a DNA virus, EBV can be sensed by DNA recognition receptors to trigger a series of downstream immune responses. A DNA-sensing pathway consists of DNA sensors, adaptor molecules and downstream effector signals. Double-stranded DNA sensors mainly include absent in melanoma 2-like receptors (ALRs) and cyclic GMP-AMP synthase (cGAS). Adaptors were mainly stimulator of interferon genes (STING) and apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC). Downstream immune responses mainly involve typeⅠIFN, inflammasomes and proinflammatory cytokines. As a double-stranded DNA virus of the Herpesviridae family, EBV triggers complex innate and adaptive immune responses in the host, especially the sensing pathways mediated by a variety of DNA recognition receptors, which play a key role in host immune defense and pathogen immune evasion. This review made the DNA sensor as the clue to comprehensively summarize the progress in the activation, regulatory mechanism and clinical relevance of DNA-sensing pathways in EBV infection in recent years, aiming to achieve a better understanding of the host innate immune responses during EBV infection and provide an immunological basis for the prevention and treatment of EBV infection-related diseases.

Hyperresponsive cytosolic DNA-sensing pathway in monocytes from primary Sjögren's syndrome.

OBJECTIVES: Cytosolic DNA-sensing pathway stimulation prompts type I IFN (IFN-I) production, but its role in systemic IFN-I pathway activation in primary SS (pSS) is poorly studied. Here we investigate the responsiveness of pSS monocytes and plasmacytoid dendritic cells (pDCs) to stimulator of interferon genes (STING) activation in relation to systemic IFN-I pathway activation and compare this with SLE. METHODS: Expression of DNA-sensing receptors cGAS, IFI16, ZBP-1 and DDX41, signalling molecules STING, TBK1 and IRF3, positive and negative STING regulators, and IFN-I-stimulated genes MxA, IFI44, IFI44L, IFIT1 and IFIT3 was analysed in whole blood, CD14+ monocytes, pDCs, and salivary glands by RT-PCR, monocyte RNA sequencing data, flow cytometry and immunohistochemical staining. Peripheral blood mononuclear cells (PBMCs) from pSS, SLE and healthy controls (HCs) were stimulated with STING agonist 2'3'-cGAMP. STING phosphorylation (pSTING) and intracellular IFNα were evaluated using flow cytometry. RESULTS: STING activation induced a significantly higher proportion of IFNα-producing monocytes, but not pDCs, in both IFN-low and IFN-high pSS compared with HC PBMCs. Additionally, a trend towards more pSTING+ monocytes was observed in pSS and SLE, most pronounced in IFN-high patients. Positive STING regulators TRIM38, TRIM56, USP18 and SENP7 were significantly higher expression in pSS than HC monocytes, while the dual-function STING regulator RNF26 was downregulated in pSS monocytes. STING was expressed in mononuclear infiltrates and ductal epithelium in pSS salivary glands. STING stimulation induced pSTING and IFNα in pSS and SLE pDCs. CONCLUSION: pSS monocytes and pDCs are hyperresponsive to stimulation of the STING pathway, which was not restricted to patients with IFN-I pathway activation.

Protective Roles of Folic Acid in the Responses of Bovine Mammary Epithelial Cells to Different Virulent Staphylococcus aureus Strains.

Mastitis caused by Staphylococcus aureus (S. aureus) infection is one of the most difficult diseases to treat in dairy cattle. Exploring the biological progression of S. aureus mastitis via the interaction between host, pathogen, and environment is the key to an effective and sustainable improvement of animal health. Here, two strains of S. aureus and a strain of MRSA (Methicillin-resistant Staphylococcus aureus) isolated from cows with different inflammation phenotypes were used to challenge Mac-T cells and to investigate their effects on the global transcriptome of the cells, then to explore the potential regulatory mechanisms of folic acid on S. aureus mastitis prevention. Differential gene expression or splicing analysis showed that different strains of S. aureus led to distinct transcriptional responses from the host immune system. Folic acid could protect host defense against the challenge of S. aureus and MRSA partially through activating cytoplasmic DNA sensing and tight junction pathway. ZBP1 at the upstream of cytoplasmic DNA sensing pathway was verified and related to anti-pathogen through RNA interference. Further enrichment analysis using these transcriptome data with cattle large-scale genome-wide association study (GWAS) data confirmed that ZBP1 gene is highly associated with bovine somatic cell score (SCS) trait. Our data shed light on the potential effect of FA through regulating key gene and then protect host cells' defense against S. aureus and MRSA.

Penaeus monodon Interferon Regulatory Factor (PmIRF) Activates IFNs and Antimicrobial Peptide Expression via a STING-Dependent DNA Sensing Pathway.

Interferon regulatory factors (IRFs) are transcription factors found in both vertebrates and invertebrates that were recently identified and found to play an important role in antiviral immunity in black tiger shrimp Penaeus monodon. In this study, we investigated the mechanism by which P. monodon IRF (PmIRF) regulates the immune-related genes downstream of the cytosolic DNA sensing pathway. Depletion of PmIRF by double-stranded RNA-mediated gene silencing significantly reduced the mRNA expression levels of the IFN-like factors PmVago1, PmVago4, and PmVago5 and antilipopolysaccharide factor 6 (ALFPm6) in shrimp. In human embryonic kidney (HEK293T) cells transfected with PmIRF or co-transfected with DEAD-box polypeptide (PmDDX41) and simulator of IFN genes (PmSTING) expression plasmids, the promoter activity of IFN-ß, nuclear factor (NF-κB), and ALFPm6 was synergistically enhanced following stimulation with the nucleic acid mimics deoxyadenylic-deoxythymidylic acid sodium salt [poly(dA:dT)] and high molecular weight (HMW) polyinosinic-polycytidylic acid [poly(I:C)]. Both nucleic acid mimics also significantly induced PmSTING, PmIRF, and ALFPm6 gene expression. Co-immunoprecipitation experiments showed that PmIRF interacted with PmSTING in cells stimulated with poly(dA:dT). PmSTING, PmIRF, and PmDDX41 were localized in the cytoplasm of unstimulated HEK293T cells and PmIRF and PmDDX41 were translocated to the nucleus upon stimulation with the nucleic acid mimics while PmSTING remained in the cytoplasm. These results indicate that PmIRF transduces the pathogen signal via the PmDDX41-PmSTING DNA sensing pathway to induce downstream production of interferon-like molecules and antimicrobial peptides.

Time-resolved RNA-seq provided a new understanding of intestinal immune response of European eel (Anguilla anguilla) following infection with Aeromonas hydrophila.

No studies systematically examined the intestinal immune response for yellow stage of European eel (Anguilla anguilla) with Aeromonas hydrophila infection by time-resolved RNA-seq. Here, we examined transcriptional profiles of the intestines at three-time points following infection with A. hydrophila. Intraperitoneal injections caused mortalities within 48 h post-injection (hpi), with the survival rate 87.5% at 24 hpi and 83.9% at 48 hpi. The result from KEGG pathway enrichment analysis showed that the immune related "cytosolic DNA-sensing pathway" was significantly enriched at the first and second time points (6 hpi and 18 hpi), with the up-regulated expression of irf3, il1b, tnfaip3, cxcl8a, ap1-2, c-fos, polr3d, polr3g and polr3k both at 6 hpi and 18 hpi, but not at the third time point (36 hpi). According to the KEGG annotation, 326 immune and inflammation-related DEGs were found. The co-expression network of those 326 DEGs revealed the existence of three modules, and tlr1 was found to be in the center of the biggest module which contained massive DEGs from "signal transduction" and "transport and catabolism". The c3 isoforms showed different expression pattern among the three time points, indicating a unique activation of complement systems at 18 hpi. Furthermore, two cathelicidins (aaCATH_1 and aaCATH_2) were highly up-regulated at the first two time points, and the bacterial growth inhibition assay revealed their antibacterial properties against A. hydrophila. Our data indicated the important roles of cytosolic DNA-sensing pathway, as well as transcripts including tlr1, c3, polr and cathelicidins in the intestine of A. anguilla in response to A. hydrophila infection. The present study will provide leads for functional studies of host-pathogen interactions.

A cytosolic sensor, PmDDX41, mediates antiviral immune response in black tiger shrimp Penaeus monodon.

DEAD (Asp-Glu-Ala-Asp)-box polypeptide 41 (DDX41), a receptor belonging to the DExD family, has recently been identified as an intracellular DNA sensor in vertebrates. Here, we report on the identification and functional characterization of PmDDX41, the first cytosolic DNA sensor in shrimp. By searching a Penaeus monodon expressed sequence tag (EST) database (http://pmonodon.biotec.or.th), three cDNA fragments exhibiting similarity to DDX41 in various species were identified and assembled, resulting in a complete open reading frame of PmDDX41 that contains 1863-bp and encodes a putative protein of 620 amino acids. PmDDX41 shares 83% and 79% similarity to DDX41 homolog from the bee Apis florea and fruit fly Drosophila melanogaster, respectively and contains three conserved domains in the protein: DEADc domain, HELICc domain, and zinc finger domain. The transcript of PmDDX41 was detected in all tested tissues and was up-regulated upon infection with a DNA virus, white spot syndrome virus (WSSV). However, PmDDX41 mRNA expression was not significantly changed and down-regulated in response to a bacterium, Vibrio harveyi, or an RNA virus, yellow head virus (YHV), respectively, compared with the control phosphate-buffered saline-injected shrimp. Furthermore, the suppression of PmDDX41 by dsRNA-mediated gene silencing resulted in more rapid death of WSSV-infected shrimp and a significant decrease in the mRNA expression levels of several immune-related genes (PmIKKß, PmIKKɛ, PmRelish, PmCactus, PmDorsal, PmPEN3, PmPEN5, and ALFPm6). These results suggest that PmDDX41 is involved in the antiviral response, probably via a DNA-sensing pathway that is triggered through the IκB kinase complex and leads to the activation of several immune-related genes.