Depletion of MicroRNA-373 Represses the Replication of Hepatitis C Virus via Activation of Type 1 Interferon Response by Targeting IRF5

PURPOSE: Hepatitis C virus (HCV) poses a risk of chronic liver disease and threatens a significant number of people worldwide. MicroRNAs (miRNAs) are linked to the regulation of hepatocarcinogenesis. Although miR-373 is required for HCV infection, the underlying mechanisms of miR-373 involvement in HCV replication remain elusive. MATERIALS AND METHODS: Quantitative reverse transcription PCR assays were performed to detect the abundances of miR-373 and HCV RNA either in Huh 7.5 cells or liver biopsy specimens with HCV infection. Luciferase assay was employed to probe the interactions between miR-373 and interferon regulatory factor 5 (IRF5). Western blot was conducted to investigate the effect of miR-373 and IRF5 on HCV replication and activation of type 1 interferon (IFN) response in JFH1-infected Huh 7.5 cells. RESULTS: HCV infection appeared to be caused by increased miR-373 expression. Addition of miR-373 promoted HCV RNA expression, while miR-373 depletion led to an inhibitive effect on HCV replication. Concordantly, IRF5, as a direct target, was limited by miR-373 in JFH1-infected Huh 7.5 cells. In addition, introduction of IRF5 protected HCV replication in the presence of abundant miR-373. Furthermore, the miR-373-mediated inhibitory effect on type 1 IFN response was ablated following IRF5 accumulation. CONCLUSION: miR-373 abrogation reduced HCV replication via activation of type 1 IFN responses by targeting IRF5 in JFH1-infected Huh 7.5 cells, suggesting a promising therapeutic for treating HCV infection.

Interferon regulatory factor 5 and systemic lupus erythematosus / 国际儿科学杂志

Interferon regulatory factor (IRF)members are composed of 10 different proteins,including IRF1 ～ IRF9 and viruses IRF(V-IRF).These IRFs regulate the transcription of type I IFN genes as transcription factors.Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by a large number of autoantibodies and precipitated immune complexes,which can cause the damage of multiple organs and systems.Type I interferon system,especially the IFN-α is an important pathogenic factor in the process of SLE morbidity.SLE patients may have high level of IFN-α,which could affect the activation of the immune system to promote the development of SLE through the regulation of a variety of immune cells' activation,differentiation and function.Besides IRF3 and IRFT,the transcription factor IRF5 gene has also been shown to be related to the production of type I interferon and is an important regulator of the IFN pathway,and its genetic polymorphism and expression abnormality lead to the susceptibility of SLE.In addition to regulating the expression of type I IFN genes,IRF5 is also associated with other signaling pathways,including B cell transformation of IgG,macrophage polarization and apoptosis,and these signaling pathways in the pathogenesis of SLE also play a very important role.This article reviews the role of IRF5 in the development of SLE disease.

Interferon Regulatory Factor 5 Inhibits Invasion Ability of Nasopharyngeal Carcinoma by Reducing PARP-1 / 中山大学学报(医学科学版)

[Objective]To investigate whether IRF5 can inhibit invasion ability of nasopharyngeal carcinoma by re-ducing PARP-1(poly(ADP-ribose)polymerase-1).[Methods]Forty-six specimens of nasopharyngeal carcinoma and 51 specimens of normal tissue were confirmed by pathologically in this study.The expression of IRF5 and PARP-1 in naso-pharyngeal carcinoma tissues and normal tissues was detected by immunohistochemistry.The IFR5 overexpression plasmid was transfected into the nasopharyngeal carcinoma cell line CNE-2,quantitative PCR and immunoblotting was used to value the expression of IRF5 after transfection.The wound healing and transwell assay was used to investigate the invasion ability. The expression of PARP-1 was valued by quantitative PCR and immunoblotting after over-expression of PFR5.[Results]The results showed that the expression of IRF5 in cancer tissues was lower than that in normal tissues,but the PARP-1 expression was opposite. The IRF5 overexpressing cell line CNE-2/IFR5 was established. The healing rate of CNE-2/IFR5 cells was lower than that of the control cells(P<0.01). Transwell experiments revealed that the number of CNE-2/IFR5 cells passing through the basement membrane was smaller than that of the control group(P<0.01),suggest-ing that up-regulation of IFR5 could inhibit the invasiveness of nasopharyngeal carcinoma cells.Over-expression of IFR5 led to reduced PARP-1 mRNA and protein(P<0.01).Besides,elevation of PARP-1 can prevent IRF5-induced changes of invasion ability.[Conclusion]Therefore,we speculated that IRF5 can inhibit invasion ability of nasopharyngeal carci-noma by reducing the expression of PARP-1.This study provided a new target for inhibiting the invasion ability of naso-pharyngeal carcinoma based on IRF5.

Effects of IRF1 on polarization and antitumor function of M 1 microphage / 基础医学与临床

Objective To study if IRF1 could regulate the polarization by IRF 1 and M1 status and affect M1 media-ted antitumor function .Methods U937 derived M1 macrophage ( U937-M1 ) model was established .The cells were devided into 4 groups:the PMA pretreated unpolarized macrophage (M0), the PMA, IFN-γand LPS induced M1 macrophage (M1), the siRNA of IRF1 knocked down M1 macrophage (siIRF1) and the negative control siR-NA treated M1 macrophage (siC).Furthermore, the expression of CD86 and CD206 was detected by flow cytome-try, the M1/M2 associated markers (IL-12p35,IL-12p40,IL-23p19,IL-6,TNF-α/IL-10) and IFNB1 were ana-lyzed by qPCR,the expression of IL-12p70 and IL-10 was examined by ELISA, the expression of IRF1 and IRF5 was detected by Western blot , the proliferration and apoptosis of HCC were analyzed by CCK 8 and flow cytometry , respectively.Results Compared with the U937-M1, the IRF1 knocked down group showed impaired CD 86 expres-sion, but enhanced CD206 expreesion ( P<0.05 ); the expression of M1 related cytokines including IL-12p35, IL-12p40,IL-23p19,IL-6,TNF-αand IFNB1 was decreased, but M2 related cytokine IL-10 level was increased (P<0.01);the expression of IFN-β, IL-12p70 and IRF5 was impaired, but IL-10 was enhanced (P<0.05).In IRF1 knocked down U937-M1, the CCK8 analysis indicated that the M1 mediated anti-proliferation effects on hepatoma carcinoma cell were turned to pro-proliferation ( P<0.05);the flow cytometry showed that the M 1 mediated pro-ap-optosis effects were reversed to anti-apoptosis ( P<0.01 ) .Interestingly , IRF5 and IFN-βwere decreased at both mRNA and protein levels in IRF1 knocked down U937-M1 compared with the U937-M1 (P<0.01).Conclusions IRF1 may partly modulate IRF5 and IFN-β, and further regulate M1 polarization and its antitumor effects .

Study on the relationship between polymorphism sites of IRF5 and SLE patients in Shandong Han population / 中华微生物学和免疫学杂志

Objective To investigate two single nucleotide polymorphism sites of IRF5 and to de-tect their relationship with SLE in a population from Shandong province. Methods The polymorphisms (rs2004640 G/T,rs10954213 G/A) were detected with PCR-RFLP in 92 eases of SLE and 88 healthy con-trols. The genotype and allele frequencies were calculated and analyzed. Results The genotype frequencies Of GG, GT and TT in rs2004640 site in SLE were 0. 198, 0.521 and 0.281, respectively. The difference was significant between SLE and centrol (X2 = 8.73, P < 0.05). The genotype frequencies of GG, GA and AA in rs10954213 site in SLE were 0. 318, 0. 409 and 0.273, respectively. The differenee was significant between SLE and control (X2 = 6. 36, P < 0. 05). Conclusion The polymorphism of rs2004640, rs10954213 in IRF5 may be associated with SLE in the population of Han nationality from Shandong province of China.

Development of Monoclonal Antibodies Against Human IRF-5 and Their Use in Identifying the Binding of IRF-5 to Nuclear Import Proteins Karyopherin-alpha1 and -beta1

PURPOSE: IRF-5 is a direct transducer of virus-mediated and TLR-mediated signaling pathways for the expression of cytokines and chemokines which form homodimers or heterodimers with IRF-7. However, direct IRF-5-specific monoclonal antibodies (mAbs) are not available at present. These could be used to further evaluate the functions of IRF-5. In this study, we produced and characterized three mouse mAbs to human IRF-5. The binding of IRF-5 to nuclear import proteins was first identified using a mAb. MATERIALS AND METHODS: His-tagged human IRF-5 protein spanning amino acid residues 193- 257 was used as an antigen and three mAbs were produced. The mAbs were tested with ELISA, Western blot analysis (WB), immunofluorescent staining (IF), and immunoprecipitation (IP). In addition, the nuclear import protein which carried phosphorylated IRF-5 was identified using one of these mAbs. RESULTS: MAbs 5IRF8, 5IRF10 and 5IRF24 which reacted with the recombinant His-IRF-5(193-257) protein were produced. All mAbs bound to human IRF-5, but not to IRF-3 or IRF-7. They could be used for WB, IF, and IP studies. The binding of phosphorylated IRF-5 to karyopherin-alpha1 and -beta1 was also identified. CONCLUSION: Human IRF-5-specific mAbs are produced for studying the immunologic roles related to IRF-5. Phosphorylated IRF-5 is transported to the nucleus by binding to nuclear import proteins karyopherin-alpha1 and -beta1.