• Biomolecules & Therapeutics
• /
• v.31 no.1
• /
• pp.48-58
• /
• 2023

Interferon regulatory factor 3 (IRF3) integrates both immunological and non-immunological inputs to control cell survival and death. Small GTPases are versatile functional switches that lie on the very upstream in signal transduction pathways, of which duration of activation is very transient. The large number of homologous proteins and the requirement for site-directed mutagenesis have hindered attempts to investigate the link between small GTPases and IRF3. Here, we constructed a constitutively active mutant expression library for small GTPase expression using Gibson assembly cloning. Small-scale screening identified multiple GTPases capable of promoting IRF3 phosphorylation. Intriguingly, 27 of 152 GTPases, including ARF1, RHEB, RHEBL1, and RAN, were found to increase IRF3 phosphorylation. Unbiased screening enabled us to investigate the sequence-activity relationship between the GTPases and IRF3. We found that the regulation of IRF3 by small GTPases was dependent on TBK1. Our work reveals the significant contribution of GTPases in IRF3 signaling and the potential role of IRF3 in GTPase function, providing a novel therapeutic approach against diseases with GTPase overexpression or active mutations, such as cancer.

• The Journal of Engineering Geology
• /
• v.3 no.1
• /
• pp.39-49
• /
• 1993

Intrinsic random function of order k(IRF-k) was used to estimate groundwater levels of nonstationaav random functions. The accuracy of IRF-k was compared to that of ordraarv krigrng assuming that the data of groundwater levels compose a stafionarv random function. Cross validation and statistical errors show that IRF-k is superior to orcinarv '(riging for the estimation of water levels. IRF-k and ordinary kriging made different contour and 3-D surface maps. The maps of IRF-k are more accurate than those of ordinary kriging.

• The Korean Journal of Physiology and Pharmacology
• /
• v.21 no.5
• /
• pp.547-554
• /
• 2017

Previous studies have demonstrated the role of hydroquinone (HQ), a hydroxylated benzene metabolite, in modulating various immune responses; however, its role in macrophage-mediated inflammatory responses is not fully understood. In this study, the role of HQ in inflammatory responses and the underlying molecular mechanism were explored in macrophages. HQ down-regulated the expression of interferon $(IFN)-{\beta}$ mRNA in LPS-stimulated RAW264.7 cells without any cytotoxicity and suppressed interferon regulatory factor (IRF)-3-mediated luciferase activity induced by TIR-domain-containing adapter-inducing interferon-${\beta}$ (TRIF) and TANK-binding kinase 1 (TBK1). A mechanism study revealed that HQ inhibited IRF-3 phosphorylation induced by lipopolysaccharide (LPS), TRIF, and AKT by suppressing phosphorylation of AKT, an upstream kinase of the IRF-3 signaling pathway. IRF-3 phosphorylation is highly induced by wild-type AKT and poorly induced by an AKT mutant, AKT C310A, which is mutated at an inhibitory target site of HQ. We also showed that HQ inhibited IRF-3 phosphorylation by targeting all three AKT isoforms (AKT1, AKT2, and AKT3) in RAW264.7 cells and suppressed IRF-3-mediated luciferase activities induced by AKT in HEK293 cells. Taken together, these results strongly suggest that HQ inhibits the production of a type I IFN, $IFN-{\beta}$, by targeting AKTs in the IRF-3 signaling pathway during macrophage-mediated inflammation.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.50 no.3
• /
• pp.302-310
• /
• 2017

Interferon regulatory factor 8 (IRF8) is essential for the development of B and T cells, as well as for the activity of dendritic cells and macrophages. We performed molecular characterization of IRF8 from rock fish, Sebastes schlegelii (Ss), and investigated the spatial and temporal profile of mRNA expression after challenge with lipopolysaccharide (LPS), polyinosinic:polycytidylic acid (poly I:C), or Streptococcus iniae. The full-length cDNA sequence of SsIRF8 was 1,657 bp, containing an ORF of 1,266 bp. The gene had a predicted molecular mass of 47.7 kDa and an isoelectric point of 5.99. The amino acid sequence coded by this gene showed the highest degree of identity (90.8%) and similarity (96.2%) with IRF8 from Oplegnathus fasciatus. The SsIRF8 mRNA was expressed ubiquitously, at varying levels, with the highest level of expression observed in the spleen. To confirm the role of SsIRF8 in mediating the immune response, we measured SsIRF8 mRNA expression in the splenic tissue at different time points after injection with LPS, poly I:C, or S. iniae. The qRT-PCR results showed that SsIRF8 mRNA expression in the poly I:C-injected group was highly upregulated 6 hr after exposure (P<0.05). Expression of SsIRF8 mRNA in the S. iniae-injected group peaked at 24 hr. These results suggest that SsIRF8 might be important in regulating the strength of the rockfish immune response to immunostimulatory agents.

• Journal of fish pathology
• /
• v.36 no.2
• /
• pp.213-228
• /
• 2023

We evaluated the transcriptional response of interferon (IFN)-related genes in rock bream iridovirus (RBIV)-infected rock bream under high-, low-, or no-mortality conditions induced by different stocking water temperatures. Under the high susceptibility condition (group A, water temperature 26℃, 100% mortality), only the Mx gene was expressed early, with prolonged expression, and with heavy viral loads of approximately 10⁶~10⁷ major capsid protein gene copies/μL from 4 to 10 days post infection (dpi). However, IRF1, IRF3, IRF8, STAT1, ISG15, PKR, Viperin, GVIN1, IFI44, and ISG56 were activated at later time points (8 dpi) and then quickly decreased (10 dpi). For the low susceptibility condition, the water temperature was set at 23℃ for 7 days (group B) and then reduced to 17℃. Group B exhibited a 28% mortality rate, in which persistent and effective antiviral responses were observed for long periods of time. In particular, at 20 and 22 dpi, when virus replication was peaked at approximately 10⁷/μL, the expressions of most of the IFN-related genes (IRF1, IRF3, IRF8, Mx, STAT1, ISG15, PKR, Viperin, GVIN1, IFI44, and ISG56) were significantly higher in group B than in the control group. Moreover, prolonged and higher levels of IRF3 (at least 30 dpi), IRF8 (at least 30 dpi), ISG15 (at least 30 dpi), PKR (at least 28 dpi), Viperin (at least 30 dpi), and IFI44 (at least 30 dpi) were also observed in the recovery stage of infection. Under the no-susceptibility condition at 17℃ (0% mortality), significantly elevated levels of IRF3, Mx, ISG15, and PKR were observed mostly until 20 dpi. The findings indicate that RBIV infection can induce an efficient IFN-mediated antiviral immune response in low- and no-susceptibility conditions. The findings could be valuable for effective control of viral pathogens in fish.

• Asian Pacific Journal of Cancer Prevention
• /
• v.14 no.11
• /
• pp.6803-6806
• /
• 2013

Background: Interferon regulatory factor 6 (IRF6) is a transcription factor with distinct and conserved DNA and protein binding domains. Mutations within the protein binding domain have been significantly observed in subjects with orofacial cleft relative to healthy controls. In addition, recent studies have identified loss of expression of IRF6 due to promoter hypermethylation in cutaneous squamous cell carcinomas. Since mutational events occurring within the conserved domains are likely to affect the function of a protein, we investigated whether regions within the IRF6 gene that encodes for the conserved protein binding domain carried mutations in oral squamous cell carcinoma (OSCC). Materials and Methods: Total chromosomal DNA extracted from 32 post surgical OSCC tissue samples were amplified using intronic primers flanking the exon 7 of IRF6 gene, which encodes for the major region of protein binding domain. The PCR amplicons from all the samples were subsequently resolved in a 1.2% agarose gel, purified and subjected to direct sequencing to screen for mutations. Results: Sequencing analysis resulted in the identification of a mutation within exon 7 of IRF6 that occurred in heterozygous condition in 9% (3/32) of OSCC samples. The wild type codon TTC at position 252 coding for phenylalanine was found to be mutated to TAC that coded for tyrosine (F252Y). Conclusions: The present study identified for the first time a novel mutation within the conserved protein binding domain of IRF6 gene in tissue samples of subjects with OSCC.

• Development and Reproduction
• /
• v.25 no.4
• /
• pp.235-244
• /
• 2021

Interferon Regulatory Factor 3 (IRF3) is a member of interferon-regulated transcription factor family and is known to play an important role in the innate immune response against viral infections. In this study, the expression of IRF3 in different tissues, developmental stages, and stocking densities of olive flounder was investigated. The expression of IRF3 was observed to gradually increase in early-stage juvenile fish. The highest expression was observed in later-stage juvenile fish when immune tissues were formed. High IRF3 expression was observed in the muscles and the brain tissues. The expression of IRF3 was observed in fish at different stocking densities after viral hemorrhagic septicemia virus (VHSV) infection. It yielded an interesting expression pattern in the muscles and the brain tissues of fish stocked at low density. These observations can be used as basic data for the study of the expression of immune response-related genes against viruses based on stocking density and immune systems in other fish species.

• Clinical and Experimental Pediatrics
• /
• v.48 no.3
• /
• pp.284-291
• /
• 2005

Purpose : Flow cytometric automated reticulocyte analysis is a superior method to manual reticulocyte counting, with respect to precision and sensitivity. Furthermore, flow cytometric analysis is able to measure immature reticulocyte fraction(IRF) and reticulocyte cellular indices(RCI : cell hemoglobin content : CHr, mean cell volume : MCVr, cell hemoglobin concentration mean : CHCMr, distribytion width : RDWr, HDWr, CHDWr). In this study, we investigated the mean values and clinical significances of IRF and RCI in healthy children and pediatric anemia patients. Methods : IRF and RCI were measured with an automated blood cell analyzer, ADVIA 120(Bayer, USA) using oxazine 750 dye, in 57 healthy children and 61 children with anemia. The anemia group consisted of 27 iron deficiency anemia(IDA) patients and 34 patients with anemia associated with acute infection(AAI). We compared the mean values of IRF and RCI in the control group classified according to age, between anemia groups and the control group, and between the IDA group and the AAI group. Results : For the normal control group, the mean values of IRF, CHr, MCVr and HDWr were higher in neonates when compared to older children. The mean values of IRF and RDWr were significantly higher, and the mean values of CHr and CHCMr were significantly lower in the IDA group when compared to the control group. The mean value of IRF was significantly higher, and the mean value of CHDWr was significantly lower in the AAI group when compared to the control group. The mean values of IRF, CHr and CHCMr were significantly lower in the IDA group when compared to the AAI group. Conclusion : We could determine the normal mean values of IRF and RCI in healthy children classified according to age for understanding of hematopoietic response differences according to age. The evaluation of IRF and RCI by automated reticulocyte analyzer seemed to be accurate and clinically useful for the early diagnosis of anemia and the differentiation of IDA from AAI.

• Korean Journal of Veterinary Research
• /
• v.44 no.2
• /
• pp.195-200
• /
• 2004

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family and potent inducer of apoptosis. TRAIL has been shown to effectively limit tumor growth in vivo without detectable cytotoxic side effects. Interferon (IFN)-${\gamma}$ often modulates the anti-cancer activities of TNF family members including TRAIL. We previously reported that IFN-${\gamma}$ enhanced TRAIL-induced Apoptosis in HeLa cells without the unknown mechanism. In this study, we investigated whether IRF-1 involves in IFN-${\gamma}$-enhanced TRAIL-induced apoptosis. We exposed HeLa cells to IFN-${\gamma}$ for 12 hours and then treated with recombinant TRAIL protein. No apoptosis was induced in cells pretreated with IFN-${\gamma}$, and TRAIL only induced 30% apoptosis after 3 hours treatment. In HeLa cells pretreated with IFN-${\gamma}$, TRAIL induced cell death to more than 75% at 3 hours, showed that IFN-${\gamma}$-pretreatment enhanced HeLa cell death to TRAIL-induced apoptosis. To investigate the functional role of IRF-1 in IFN-${\gamma}$-enhanced TRAIL-induced apoptosis, IRF-1 was overexpressed by using an adenoviral vector AdIRF-1. IRF-1 overexpression increased apoptotic cell death and significantly enhanced apoptotic cell death induced by TRAIL when infected cells were treated with TRAIL. Our findings show that IFN-${\gamma}$ enhances TRAIL-induced apoptosis by IRF-1 in HeLa cells.

• Journal of fish pathology
• /
• v.34 no.1
• /
• pp.23-29
• /
• 2021

Interferon regulatory factors (IRFs) are a family of transcription factors essential to the control of antiviral immune response, cell growth, differentiation and apoptosis. IRF10 of zebrafish (Danio rerio) was negative regulation of the interferonΦ1 and 3 response in vitro. In this study, we analyze the induction of in vivo immune response activation from the IRF10 gene of zebrafish and the protective effect against VHSV. As the results, the group inoculated with IRF10 expression vectors, there was no expression of IFNΦ1, suggestion that IRF10 may function as a negative regulator of IRF3, which binds to the IFNΦ1 promoter. And other types of interferon genes (IFNΦ2-4) are thought to have been activated, inducing to the expression of pro-inflammatory cytokine and Mx genes. As the results of challenge test performed at 14 days after inoculation of the expression vectors, the maximum survival rate [50% (1㎍ DNA) and 42.5% (10㎍ DNA)] for IRF10 group were recorded. Meanwhile, the survival rates of pcDNA3.1 and PBS as the control groups were 10% and 15%, respectively. This study suggests that the possibility that activation of IRF10 molecule could be exploited as a VHS control method.