Ultraviolet-C light at 222 nm has a high disinfecting spectrum in environments contaminated by infectious pathogens, including SARS-CoV-2.

Ultraviolet light (UV) acts as a powerful disinfectant and can prevent contamination of personal hygiene from various contaminated environments. The 222-nm wavelength of UV-C has a highly effective sterilization activity and is safer than 275-nm UV-C. We investigated the irradiation efficacy of 222-nm UV-C against contaminating bacteria and viruses in liquid and fabric environments. We conducted colony-forming unit assays to determine the number of viable cells and a 50% tissue culture infectious dose assay to evaluate the virus titration. A minimum dose of 27 mJ/cm2 of 222-nm UV-C was required for >95% germicidal activity for gram-negative and -positive bacteria. A 25.1 mJ/cm2 dose could ensure >95% virucidal activity against low-pathogenic avian influenza virus and severe acute respiratory syndrome coronavirus (SARS-CoV-2). In addition, this energy dose of 222-nm UV-C effectively inactivated SARS-CoV-2 variants, Delta and Omicron. These results provide valuable information on the disinfection efficiency of 222-nm UV-C in bacterial and virus-contaminated environments and can also develop into a powerful tool for individual hygiene.

Pseudoknot-targeting Cas13b combats SARS-CoV-2 infection by suppressing viral replication.

CRISPR-Cas13-mediated viral genome targeting is a novel strategy for defending against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants. Here, we generated mRNA-encoded Cas13b targeting the open reading frame 1b (ORF1b) region to effectively degrade the RNA-dependent RNA polymerase gene. Of the 12 designed CRISPR RNAs (crRNAs), those targeting the pseudoknot site upstream of ORF1b were found to be the most effective in suppressing SARS-CoV-2 propagation. Pseudoknot-targeting Cas13b reduced expression of the spike protein and attenuated viral replication by 99%. It also inhibited the replication of multiple SARS-CoV-2 variants, exhibiting broad potency. We validated the therapeutic efficacy of this system in SARS-CoV-2-infected hACE2 transgenic mice, demonstrating that crRNA treatment significantly reduced viral titers. Our findings suggest that the pseudoknot region is a strategic site for targeted genomic degradation of SARS-CoV-2. Hence, pseudoknot-targeting Cas13b could be a breakthrough therapy for overcoming infections by SARS-CoV-2 or other RNA viruses.

Delta (B1.617.2) variant of SARS-CoV-2 induces severe neurotropic patterns in K18-hACE2 mice.

A highly contagious virus, severe acute respiratory syndrome coronavirus 2, caused the coronavirus disease 19 (COVID-19) pandemic (SARS-CoV-2). SARS-CoV-2 genetic variants have been reported to circulate throughout the COVID-19 pandemic. COVID-19 symptoms include respiratory symptoms, fever, muscle pain, and breathing difficulty. In addition, up to 30% of COVID-19 patients experience neurological complications such as headaches, nausea, stroke, and anosmia. However, the neurotropism of SARS-CoV-2 infection remains largely unknown. This study investigated the neurotropic patterns between the B1.617.2 (Delta) and Hu-1 variants (Wuhan, early strain) in K18-hACE2 mice. Despite both the variants inducing similar pathogenic patterns in various organs, B1.617.2-infected K18-hACE2 mice demonstrated a higher range of disease phenotypes such as weight loss, lethality, and conjunctivitis when compared to those in Hu-1-infected mice. In addition, histopathological analysis revealed that B1.617.2 infects the brain of K18-hACE2 mice more rapidly and effectively than Hu-1. Finally, we discovered that, in B1.617.2-infected mice, the early activation of various signature genes involved innate cytokines and that the necrosis-related response was most pronounced than that in Hu-1-infected mice. The present findings indicate the neuroinvasive properties of SARS-CoV-2 variants in K18-hACE2 mice and link them to fatal neuro-dissemination during the disease onset.

Hepatitis C Virus Nonstructural Protein 5A Interacts with Immunomodulatory Kinase IKKε to Negatively Regulate Innate Antiviral Immunity.

Hepatitis C virus (HCV) infection can lead to chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. HCV employs diverse strategies to evade host antiviral innate immune responses to mediate a persistent infection. In the present study, we show that nonstructural protein 5A (NS5A) interacts with an NF-κB inhibitor immunomodulatory kinase, IKKε, and subsequently downregulats beta interferon (IFN-ß) promoter activity. We further demonstrate that NS5A inhibits DDX3-mediated IKKε and interferon regulatory factor 3 (IRF3) phosphorylation. We also note that hyperphosphorylation of NS5A mediats protein interplay between NS5A and IKKε, thereby contributing to NS5A-mediated modulation of IFN-ß signaling. Lastly, NS5A inhibits IKKε-dependent p65 phosphorylation and NF-κB activation. Based on these findings, we propose NS5A as a novel regulator of IFN signaling events, specifically by inhibiting IKKε downstream signaling cascades through its interaction with IKKε. Taken together, these data suggest an additional mechanistic means by which HCV modulates host antiviral innate immune responses to promote persistent viral infection.

Evaluation of Antiviral Effect against SARS-CoV-2 Propagation by Crude Polysaccharides from Seaweed and Abalone Viscera In Vitro.

Crude polysaccharides, extracted from two seaweed species (Hizikia fusiforme and Sargassum horneri) and Haliotis discus hannai (abalone) viscera, were evaluated for their inhibitory effect against SARS-CoV-2 propagation. Plaque titration revealed that these crude polysaccharides efficiently inhibited SARS-CoV-2 propagation with IC50 values ranging from 0.35 to 4.37 µg/mL. The crude polysaccharide of H. fusiforme showed the strongest antiviral effect, with IC50 of 0.35 µg/mL, followed by S. horneri and abalone viscera with IC50 of 0.56 and 4.37 µg/mL, respectively. In addition, immunofluorescence assay, western blot, and quantitative RT-PCR analysis verified that these polysaccharides could inhibit SARS-CoV-2 replication. In Vero E6 cells, treatment with these crude polysaccharides before or after viral infection strongly inhibited the expression level of SARS-CoV-2 spikes, nucleocapsid proteins, and RNA copies of RNA-dependent RNA-polymerase and nucleocapsid. These results show that these crude marine polysaccharides effectively inhibit SARS-CoV-2 propagation by interference with viral entry.

Hepatitis C Virus Nonstructural 5A Protein Interacts with Telomere Length Regulation Protein: Implications for Telomere Shortening in Patients Infected with HCV.

Hepatitis C virus (HCV) is a major cause of chronic liver disease and is highly dependent on cellular proteins for viral propagation. Using protein microarray analysis, we identified 90 cellular proteins as HCV nonstructural 5A (NS5A) interacting partners, and selected telomere length regulation protein (TEN1) for further study. TEN1 forms a heterotrimeric complex with CTC and STN1, which is essential for telomere protection and maintenance. Telomere length decreases in patients with active HCV, chronic liver disease, and hepatocellular carcinoma. However, the molecular mechanism of telomere length shortening in HCV-associated disease is largely unknown. In the present study, protein interactions between NS5A and TEN1 were confirmed by immunoprecipitation assays. Silencing of TEN1 reduced both viral RNA and protein expression levels of HCV, while ectopic expression of the siRNA-resistant TEN1 recovered the viral protein level, suggesting that TEN1 was specifically required for HCV propagation. Importantly, we found that TEN1 is re-localized from the nucleus to the cytoplasm in HCV-infected cells. These data suggest that HCV exploits TEN1 to promote viral propagation and that telomere protection is compromised in HCV-infected cells. Overall, our findings provide mechanistic insight into the telomere shortening in HCV-infected cells.

Adenosylhomocysteinase like 1 interacts with nonstructural 5A and regulates hepatitis C virus propagation.

Hepatitis C virus (HCV) life cycle is highly dependent on cellular proteins for viral propagation. In order to identify the cellular factors involved in HCV propagation, we previously performed a protein microarray assay using the HCV nonstructural 5A (NS5A) protein as a probe. Of ∼9,000 human cellular proteins immobilized in a microarray, adenosylhomocysteinase like 1 (AHCYL1) was among 90 proteins identified as NS5A interactors. Of these candidates, AHCYL1 was selected for further study. In the present study, we verified the physical interaction between NS5A and AHCYL1 by both in vitro pulldown and coimmunoprecipitation assays. Furthermore, HCV NS5A interacted with endogenous AHCYL1 in Jc1-infected cells. Both NS5A and AHCYL1 were colocalized in the cytoplasmic region in HCV-replicating cells. siRNAmediated knockdown of AHCYL1 abrogated HCV propagation. Exogenous expression of the siRNA-resistant AHCYL1 mutant, but not of the wild-type AHCYL1, restored HCV protein expression levels, indicating that AHCYL1 was required specifically for HCV propagation. Importantly, AHCYL1 was involved in the HCV internal ribosome entry site-mediated translation step of the HCV life cycle. Finally, we demonstrated that the proteasomal degradation pathway of AHCYL1 was modulated by persistent HCV infection. Collectively, these data suggest that HCV may modulate the AHCYL1 protein to promote viral propagation.

Cortactin Interacts with Hepatitis C Virus Core and NS5A Proteins: Implications for Virion Assembly.

Hepatitis C virus (HCV) exploits cellular proteins to facilitate viral propagation. To identify the cellular factors involved in the HCV life cycle, we previously performed protein microarray assays using either HCV nonstructural 5A (NS5A) protein or core protein as a probe. Interestingly, cellular cortactin strongly interacted with both NS5A and core. Cortactin is an actin-binding protein critically involved in tumor progression by regulating the migration and invasion of cancerous cells. Protein interaction between cortactin and NS5A or core was confirmed by coimmunoprecipitation and immunofluorescence assays. We showed that cortactin interacted with NS5A and core via the N-terminal acidic domain of cortactin. Cortactin expression levels were not altered by HCV infection. Small interfering RNA (siRNA)-mediated knockdown of cortactin dramatically decreased HCV protein expression and infectivity levels, whereas overexpression of cortactin increased viral propagation. Ectopic expression of the siRNA-resistant cortactin recovered the viral infectivity, suggesting that cortactin was specifically required for HCV propagation. We further showed that cortactin was involved in the assembly step without affecting viral entry, HCV internal ribosome entry site (IRES)-mediated translation, and the replication steps of the HCV life cycle. Of note, silencing of cortactin markedly reduced both NS5A and core protein levels on the lipid droplets (LDs), and this effect was reversed by the overexpression of cortactin. Importantly, NS5A and core promoted cell migration by activating the phosphorylation of cortactin at tyrosine residues 421 and 466. Taken together, these data suggest that cortactin is not only involved in HCV assembly but also plays an important role in the cell migration.IMPORTANCE Cortactin is a cytoskeletal protein that regulates cell migration in response to a number of extracellular stimuli. The functional involvement of cortactin in the virus life cycle is not yet fully understood. The most significant finding is that cortactin strongly interacted with both hepatitis C virus (HCV) core and NS5A. Cortactin is involved in HCV assembly by tethering core and NS5A on the lipid droplets (LDs) with no effect on LD biogenesis. It was noteworthy that HCV NS5A and core activated cortactin by phosphorylation at tyrosines 421 and 466 to regulate cell migration. Collectively, our study shows that cortactin is a novel host factor involved in viral production and HCV-associated pathogenesis.

Nonstructural NS5A Protein Regulates LIM and SH3 Domain Protein 1 to Promote Hepatitis C Virus Propagation.

Hepatitis C virus (HCV) propagation is highly dependent on cellular proteins. To identify the host factors involved in HCV propagation, we previously performed protein microarray assays and identified the LIM and SH3 domain protein 1 (LASP-1) as an HCV NS5A-interacting partner. LASP-1 plays an important role in the regulation of cell proliferation, migration, and protein-protein interactions. Alteration of LASP-1 expression has been implicated in hepatocellular carcinoma. However, the functional involvement of LASP1 in HCV propagation and HCV-induced pathogenesis has not been elucidated. Here, we first verified the protein interaction of NS5A and LASP-1 by both in vitro pulldown and coimmunoprecipitation assays. We further showed that NS5A and LASP-1 were colocalized in the cytoplasm of HCV infected cells. NS5A interacted with LASP-1 through the proline motif in domain I of NS5A and the tryptophan residue in the SH3 domain of LASP-1. Knockdown of LASP-1 increased HCV replication in both HCV-infected cells and HCV subgenomic replicon cells. LASP-1 negatively regulated viral propagation and thereby overexpression of LASP-1 decreased HCV replication. Moreover, HCV propagation was decreased by wild-type LASP-1 but not by an NS5A binding-defective mutant of LASP-1. We further demonstrated that LASP-1 was involved in the replication stage of the HCV life cycle. Importantly, LASP-1 expression levels were increased in persistently infected cells with HCV. These data suggest that HCV modulates LASP-1 via NS5A in order to regulate virion levels and maintain a persistent infection.

Comparison of clinical outcomes between single and double vitrified-warmed blastocyst embryo transfer according to the day of vitrification.

PURPOSE: To compare the efficacy of single vitrified-warmed blastocyst embryo transfer (SVBT) versus double vitrified-warmed blastocyst embryo transfer (DVBT) according to the day of vitrification. METHODS: This retrospective study included a total of 1,051 cycles in women less than 37 years of age with their autologous SVBT cryopreserved on day 5 (5d-SVBT, n = 737) or day 6 (6d-SVBT, n = 154) and DVBT on day 5 (5d-DVBT, n = 129) or day 6 (6d-DVBT, n = 31) from January 2009 to December 2011. RESULTS: The clinical pregnancy rate (41.8 % vs. 48.1 %, p = 0.184) and ongoing pregnancy rate (36.6 % vs. 45.0 %, p = 0.072) were not significantly different between the 5d-SVBT group and the 5d-DVBT group. However, the clinical pregnancy (29.9 % vs. 58.1 %, p = 0.003) and ongoing pregnancy rates (23.4 % vs. 51.6 %, p = 0.001) were significantly lower in the 6d-SVBT group compared with those in the 6d-DVBT group. The implantation rate (42.2 % vs. 34.5 %, p = 0.03) of the 5d-SVBT group was significantly higher than that of the 5d-DVBT group, while the implantation rate (29.9 % vs. 37.1 %, p = 0.303) of the 6d-SVBT group was not statistically different compared with that in the 6d-DVBT group. The multiple pregnancy rates (1.0 % in the 5d-SVBT group vs. 38.7 % in the 5d-DVBT group, p < 0.001 and 0 % in the 6d-SVBT group vs. 22.2 % in the 6d-DVBT group, p = 0.001) were statistically significantly lower in the SVBT group compared with those in the DVBT group regardless of the day of vitrification. CONCLUSIONS: This study showed that the 5d-SVBT resulted in comparable clinical outcomes compared to the 5d-DVBT while the 6d-SVBT yielded significantly lower clinical outcomes compared to the 6d-DVBT.

Comparison of lateral opening wedge calcaneal osteotomy and medial calcaneal sliding-opening wedge cuboid-closing wedge cuneiform osteotomy for correction of planovalgus foot deformity in children.

The purpose of the present study was to compare the clinical and radiographic results between 2 procedures, lateral opening wedge calcaneal osteotomy (LCL) and medial calcaneal sliding-opening wedge cuboid-closing wedge cuneiform osteotomy (3C) in patients with planovalgus foot deformity. A total of 38 patients who underwent either LCL (18 patients, 28 feet) or 3C (20 patients, 32 feet) were included in the present study. The etiology of the planovalgus foot deformity was idiopathic in 16 feet and cerebral palsy in 44 feet. The 2 procedures used in the present study were indicated in symptomatic (pain or callus) children in whom conservative treatment, such as shoe modifications or orthotics, had been applied for more than 1 year but had failed. The patients were evaluated preoperatively, postoperatively, and at the last follow-up visit, both clinically and radiologically, and the interval to union and postoperative courses were compared between the 2 groups. In the LCL group, 19 of the 28 feet (68%) showed a satisfactory outcome and 9 (32%) an unsatisfactory outcome. In the 3C group, 28 of the 32 feet (88%) showed a satisfactory outcome and 4 (12%) an unsatisfactory outcome. The clinical results were not significantly different between the 2 groups, with mild to moderate pes planovalgus deformity. However, the clinical results were better in the 3C group with severe pes planovalgus deformity than in the LCL group with severe pes planovalgus deformity. All 4 radiographic parameters were improved at the last follow-up visit in both groups. In particular, the talar-first metatarsal angle and the calcaneal pitch angle on the weightbearing lateral radiographs were significantly improved in the 3C group with mild to moderate planovalgus foot deformity. All 4 parameters were significantly improved in the 3C group with severe planovalgus foot deformity. No significant differences were observed between the 2 groups in terms of the interval to union and postoperative care. No case of postoperative deep infection or nonunion was encountered in either group. 3C is a more effective procedure than LCL for the correction of pes planovalgus deformity in children, especially severe pes planovalgus deformities.

Saponin inhibits hepatitis C virus propagation by up-regulating suppressor of cytokine signaling 2.

Saponins are a group of naturally occurring plant glycosides which possess a wide range of pharmacological properties, including anti-tumorigenic and antiviral activities. To investigate whether saponin has anti-hepatitis C virus (HCV) activity, we examined the effect of saponin on HCV replication. HCV replication was efficiently inhibited at a concentration of 10 µg/ml of saponin in cell culture grown HCV (HCVcc)-infected cells. Inhibitory effect of saponin on HCV replication was verified by quantitative real-time PCR, reporter assay, and immunoblot analysis. In addition, saponin potentiated IFN-α-induced anti-HCV activity. Moreover, saponin exerted antiviral activity even in IFN-α resistant mutant HCVcc-infected cells. To investigate how cellular genes were regulated by saponin, we performed microarray analysis using HCVcc-infected cells. We demonstrated that suppressor of cytokine signaling 2 (SOCS2) protein level was distinctively increased by saponin, which in turn resulted in inhibition of HCV replication. We further showed that silencing of SOCS2 resurrected HCV replication and overexpression of SOCS2 suppressed HCV replication. These data imply that saponin inhibits HCV replication via SOCS2 signaling pathway. These findings suggest that saponin may be a potent therapeutic agent for HCV patients.

Clinical outcomes of elective single morula embryo transfer versus elective single blastocyst embryo transfer in IVF-ET.

PURPOSE: To compare the clinical outcomes of elective single morula embryo transfer (eSMET) versus elective single blastocyst embryo transfer (eSBET) in selected patients. METHODS: This study was a retrospective study which analyzed for 271 cycles in women under 37 years of age who are undergoing their first or second trial of in vitro fertilization-embryo transfer (IVF-ET) from January 2008 to December 2009. The eSMET was performed on day 4 (n = 130) and the eSBET was conducted on day 5 (n = 141). RESULTS: The clinical pregnancy rate (51.5% vs. 51.8%, p = 0.97), implantation rate (52.3% vs. 52.5%, p = 0.98), and live birth rate (39.2% vs. 44.7%, p = 0.36) were similar in the eSMET and eSBET groups, respectively. The miscarriage rate of the eSMET group (23.9%) was slightly higher than that of the eSBET group (13.7%) (p = 0.12), without reaching statistical significance. There was only one case of monozygotic twin pregnancy in each group. CONCLUSIONS: The clinical outcomes of day 4 eSMET were comparable to those of day 5 eSBET. Therefore, day 4 eSMET is a viable option or an alternative to day 5 eSBET, with no difference in success rates.

c-Fos regulates hepatitis C virus propagation.

Hepatitis C virus (HCV) RNA replication requires cellular factors as well as viral non-structural proteins (NS protein). Using small interfering RNA (siRNA) library screening, we previously identified c-Fos as a host factor involved in HCV propagation. In the present study, we demonstrated that silencing of c-Fos expression resulted in decrease of HCV propagation in cell culture grown HCV (HCVcc)-infected cells; whereas overexpression of c-Fos significantly increased HCV propagation. We further confirmed the positive role of c-Fos in HCV propagation by both HCV-luciferase reporter assay and immunofluorescence analysis. We showed that c-Fos level was upregulated by HCV infection. Furthermore, phorbol 12-myristate 13-acetate (PMA)-induced c-Fos level was synergistically increased by HCV infection. These data suggest that c-Fos acts as a positive regulator of HCV propagation and may contribute to HCV-associated pathogenesis.

Comparison of elective single cleavage-embryo transfer to elective single blastocyst-embryo transfer in human IVF-ET.

OBJECTIVE: This study was carried out to compare the clinical outcome of elective single cleavage-embryo transfer (eSCET) to that of elective single blastocyst-embryo transfer (eSBET) in human IVF-ET. METHODS: This study was a retrospective study which analyzed for 614 women who visited the Daegu Maria Clinic from August 2008 to December 2009. All were under 37 years old and had more than 8 mm of endometrial thickness on the day of hCG administration and at least one good quality embryo on day 3. The eSCETs were performed on day 3 (n=450) and the eSBETs were conducted on day 5 (n=164). RESULTS: The numbers of retrieved oocytes, fertilized oocytes, and day 3 good quality embryos were significantly lower in the eSCET group (12.1±6.0, 8.2±4.6, and 4.2±3.1, respectively) compared to the eSBET group (16.7±7.2, 12.1±5.0, and 8.5±4.5, respectively; p<0.001). However, the clinical pregnancy, implantation, on-going pregnancy, and live birth rates of the eSCET group (46.7, 46.9, 40.0, and 36.7%, respectively) were not statistically different from those of the eSBET group (51.2, 51.8, 45.1, and 43.9%, respectively; p=0.318, 0.278, 0.254, and 0.103, respectively). CONCLUSION: These results suggested that elective single embryo transfer should be performed regardless of the developmental stage to women less than 37 years old who had more than 8 mm of endometrial thickness on the hCG administration day and at least one good quality embryo on day 3 in order to reduce the twin pregnancy rate without reducing the whole pregnancy rate.

Modulation of interferon signaling by hepatitis C virus non-structural 5A protein: implication of genotypic difference in interferon treatment.

Interferon (IFN) response rate in hepatitis C virus (HCV) patients has been varied with genotypes. In this study, we investigated the effects of HCV NS5A protein on IFN resistance and compared the genotypic differences of NS5A. We showed that IFN-α-, poly I:C-, and Sendai virus-induced ISRE transcriptional activities were inhibited by both genotype 1b and 2a NS5A protein. We demonstrated that not only genotype 1b but also genotype 2a NS5A exerted the similar extent of IFN-α-induced antiviral activity. We showed that NS5A derived from both genotype 1b and 2a showed no significant differential IFN responses as seen in HCV patients. These data imply that some other host factor may be involved in genotypic differences of IFN antagonism in HCV patients.

The gene for aromatase, a rate-limiting enzyme for local estrogen biosynthesis, is a downstream target gene of Runx2 in skeletal tissues.

The essential osteoblast-related transcription factor Runx2 and the female steroid hormone estrogen are known to play pivotal roles in bone homeostasis; however, the functional interaction between Runx2- and estrogen-mediated signaling in skeletal tissues is minimally understood. Here we provide evidence that aromatase (CYP19), a rate-limiting enzyme responsible for estrogen biosynthesis in mammals, is transcriptionally regulated by Runx2. Consistent with the presence of multiple Runx2 binding sites, the binding of Runx2 to the aromatase promoter was demonstrated in vitro and confirmed in vivo by chromatin immunoprecipitation assays. The bone-specific aromatase promoter is activated by Runx2, and endogenous aromatase gene expression is upregulated by Runx2 overexpression, establishing the aromatase gene as a target of Runx2. The biological significance of the Runx2 transcriptional control of the aromatase gene is reflected by the enhanced estrogen biosynthesis in response to Runx2 in cultured cells. Reduced in vivo expression of skeletal aromatase gene and low bone mineral density are evident in Runx2 mutant mice. Collectively, these findings uncover a novel link between Runx2-mediated osteoblastogenic processes and the osteoblast-mediated biosynthesis of estrogen as an osteoprotective steroid hormone.

Nonstructural 5A protein activates beta-catenin signaling cascades: implication of hepatitis C virus-induced liver pathogenesis.

BACKGROUND/AIMS: The nonstructural 5A (NS5A) protein of hepatitis C virus (HCV) has been implicated in HCV-induced liver pathogenesis. Wnt/beta-catenin signaling has also been involved in tumorigenesis. To elucidate the molecular mechanism of HCV pathogenesis, we examined the potential effects of HCV NS5A protein on Wnt/beta-catenin signal transduction cascades. METHODS: The effects of NS5A protein on beta-catenin signaling cascades in hepatic cells were investigated by luciferase reporter gene assay, confocal microscopy, immunoprecipitation assay, and immunoblot analysis. RESULTS: beta-Catenin-mediated transcriptional activity is elevated by NS5A protein, in the context of HCV replication, and by infection of cell culture-produced HCV. NS5A protein directly interacts with endogenous beta-catenin and colocalizes with beta-catenin in the cytoplasm. NS5A protein inactivates glycogen synthase kinase 3beta and increases subsequent accumulation of beta-catenin in HepG2 cells. beta-Catenin was also accumulated in HCV patients' liver tissues. In addition, the accumulation of beta-catenin in HCV replicon cells requires both activation of phosphatidylinositol 3-kinase and inactivation of GSK3beta. CONCLUSIONS: NS5A activates beta-catenin signaling cascades through increasing the stability of beta-catenin. This modulation is accomplished by the protein interplay between viral and cellular signaling transducer. These data suggest that NS5A protein may directly be involved in Wnt/beta-catenin-mediated liver pathogenesis.

Hepatitis delta virus large antigen sensitizes to TNF-alpha-induced NF-kappaB signaling.

Hepatitis delta virus (HDV) infection causes fulminant hepatitis and liver cirrhosis. To elucidate the molecular mechanism of HDV pathogenesis, we examined the effects of HDV viral proteins, the small hepatitis delta antigen (SHDAg) and the large hepatitis delta antigen (LHDAg), on NF-kappaB signaling pathway. In this study, we demonstrated that TNF-alpha-induced NF-kappaB transcriptional activation was increased by LHDAg but not by SHDAg in both HEK293 and Huh7 cells. Furthermore, LHDAg promoted TRAF2-induced NF-kappaB activation. Using coimmunoprecipitation assays, we demonstrated that both SHDAg and LHDAg interacted with TRAF2 protein. We showed that isoprenylation of LHDAg was not required for the increase of NF-kappaB activity. We further showed that only LHDAg but not SHDAg increased the TNF-alpha-mediated nuclear translocation of p65. This was accomplished by activation of IkappaBalpha degradation by LHDAg. Finally, we demonstrated that LHDAg augmented the COX-2 expression level in Huh7 cells. These data suggest that LHDAg modulates NF-kappaB signaling pathway and may contribute to HDV pathogenesis.