Novel role of LLGL2 silencing in autophagy: reversing epithelial-mesenchymal transition in prostate cancer.

PURPOSE: Prostate cancer (PCa) is a major urological disease that is associated with significant morbidity and mortality in men. LLGL2 is the mammalian homolog of Lgl. It acts as a tumor suppressor in breast and hepatic cancer. However, the role of LLGL2 and the underlying mechanisms in PCa have not yet been elucidated. Here, we investigate the role of LLGL2 in the regulation of epithelial-mesenchymal transition (EMT) in PCa through autophagy in vitro and in vivo. METHODS: PC3 cells were transfected with siLLGL2 or plasmid LLGL2 and autophagy was examined. Invasion, migration, and wound healing were assessed in PC3 cells under autophagy regulation. Tumor growth was evaluated using a shLLGL2 xenograft mouse model. RESULTS: In patients with PCa, LLGL2 levels were higher with defective autophagy and increased EMT. Our results showed that the knockdown of LLGL2 induced autophagy flux by upregulating Vps34 and ATG14L. LLGL2 knockdown inhibits EMT by upregulating E-cadherin and downregulating fibronectin and α-SMA. The pharmacological activation of autophagy by rapamycin suppressed EMT, and these effects were reversed by 3-methyladenine treatment. Interestingly, in a shLLGL2 xenograft mouse model, tumor size and EMT were decreased, which were improved by autophagy induction and worsened by autophagy inhibition. CONCLUSION: Defective expression of LLGL2 leads to attenuation of EMT due to the upregulation of autophagy flux in PCa. Our results suggest that LLGL2 is a novel target for alleviating PCa via the regulation of autophagy.

Peripheral zone thickness in preoperative MRI is predictive of Trifecta achievement after Holmium laser enucleation of the prostate (HoLEP).

PURPOSE: To investigate various anatomical features of the prostate using preoperative MRI and patients' clinical factors to identify predictors of successful Holmium:YAG laser enucleation of the prostate (HoLEP). METHODS: 71 patients who had received HoLEP and undergone a 3.0-T prostate MRI scan within 6 months before surgery were retrospectively enrolled. MRI features (e.g., total prostate and transitional zone volume, peripheral zone thickness [PZT], BPH patterns, prostatic urethral angle, intravesical prostatic protrusion, etc.) and clinical data (e.g., age, body mass index, surgical technique, etc.) were analyzed using univariable and multivariable logistic regression to identify predictors of successful HoLEP. Successful HoLEP was defined as achieving the Trifecta, characterized by the contemporary absence of postoperative complications within 3 months, a 3-month postoperative maximum flow rate (Qmax) > 15 mL/s, and no urinary incontinence at 3 months postoperatively. RESULTS: Trifecta achievement at 3 months post-surgery was observed in 37 (52%) patients. Patients with Trifecta achievement exhibited a lower preoperative IPSS-quality of life score (QoL) (4.1 vs. 4.5, P = 0.016) and a thinner preoperative peripheral zone thickness (PZT) on MRI (7.9 vs.10.3 mm, P < 0.001). In the multivariable regression analysis, a preoperative IPSS-QoL score < 5 (OR 3.98; 95% CI, 1.21-13.07; P = 0.017) and PZT < 9 mm (OR 11.51; 95% CI, 3.51-37.74; P < 0.001) were significant predictors of Trifecta achievement after HoLEP. CONCLUSIONS: Alongside the preoperative QoL score, PZT measurement in prostate MRI can serve as an objective predictor of successful HoLEP. Our results underscore an additional utility of prostate MRI beyond its role in excluding concurrent prostate cancer.

Single-port robot-assisted pyeloplasty using the da vinci SP system versus multiport pyeloplasty: Comparison of outcomes and costs.

OBJECTIVE: To investigate the feasibility of single-port (SP) robotic pyeloplasty by comparing perioperative outcomes with those of multiport (MP) robotic pyeloplasty. MATERIALS AND METHODS: We reviewed the data from patients who underwent robot-assisted pyeloplasty for ureteropelvic junction obstruction (UPJO) at a single tertiary institution between March 2016 and May 2022. Radiographic and symptomatic improvements were assessed 3 months postoperatively. Propensity score matching was performed for age, sex, body mass index, and hydronephrosis grade. RESULTS: Of the 15 S P-pyeloplasty and 28 MP-pyeloplasty cases, 14 from each group were matched using 1:1 matching. The SP group had shorter console and operative times without significant differences. Blood loss was lower in the SP group than in the MP group (p = 0.019). The length of hospital stay, opioid use on the operative day, and pain score at discharge did not differ between the two groups. The mean cost for surgery was higher in the SP group than in the MP group (p < 0.001). The mean cost of hospitalization was comparable between the two groups (p = 0.083). The cosmetic numerical rating scale scores were significantly higher in the SP group (p = 0.014). Symptoms improved in all patients, and the radiographic improvement rates were 92.9% in the SP group and 100% in the MP group. CONCLUSION: SP-pyeloplasty showed cosmetic benefits, lower blood loss, operative time, and console time compared with MP-pyeloplasty. In patients who underwent surgery for UPJO for the first time, SP surgery can show comparable outcomes when compared to MP surgery.

Clinical Validation of the Proenkephalin (PENK) Methylation Urine Test for Monitoring Recurrence of Non-muscle-invasive Bladder Cancer.

Background and objective: To assess the effectiveness of a urine-based proenkephalin (PENK) methylation test using linear target enrichment-quantitative methylation-specific polymerase chain reaction (mePENK test) for detection of non-muscle-invasive bladder cancer (NMIBC) recurrence compared to cytology and the NMP22 test. Methods: We first conducted a retrospective case-control study involving 54 patients with primary BC and 29 healthy individuals. We then prospectively enrolled 186 patients (January to December 2022) undergoing cystoscopy surveillance after transurethral resection of bladder tumor, of whom 59 had recurrent tumors. We analyzed voided urine samples for PENK methylation levels in urinary DNA. Cystoscopy with histology was used as the reference standard for assessing the diagnostic accuracy of the mePENK test in detecting BC recurrence. We calculated the sensitivity and specificity using receiver operating characteristic curve analysis. Survival differences were determined using the Kaplan-Meier method and Cox proportional-hazards model. A p < 0.05 was considered statistically significant. Key findings and limitations: In the case-control study, the PENK test had sensitivity of 83.3% and specificity of 100%. For NMIBC patients undergoing cystoscopy surveillance, the sensitivity was 76.3% (95% confidence interval [CI] 63.4-86.4%) and the specificity was 85% (95% CI 77.6-90.7%), outperforming cytology (sensitivity: 28.8%, 95% CI 17.8-42.1%; p < 0.001; specificity: 97.6%, 95% CI 93.2-99.5%) and the NMP22 test (sensitivity: 54.2%, 95% CI 40.7-67.2%; p = 0.016; specificity 81.9%, 95% CI 74.1-88.2%). In the high-risk group, the mePENK test had sensitivity of 89.7% (95% CI 75.8-97.1%) and a negative predictive value of 96.9%. For the group with low/intermediate risk, the sensitivity was 41.7%. In the group with negative cystoscopy, recurrence-free survival was shorter for patients with positive than for those with negative mePENK results (245 vs 503 d), with a hazard ratio of 9.4 (p < 0.001). The main study limitation is the small sample size. Conclusions and clinical implications: The mePENK test showed good performance for detection of NMIBC recurrence and has potential for use for prognosis and prediction. Patient summary: We found that a test used to analyze urine samples showed good performance in detecting recurrence of NMIBC. This noninvasive mePENK test may help in personalized follow-up care for patients with NMIBC.

SIAH1 modulates antiviral immune responses by targeting deubiquitinase USP19.

Tight control of the type I interferon (IFN) signaling pathway is critical for maintaining host innate immune responses, and the ubiquitination and deubiquitination of signaling molecules are essential for signal transduction. Deubiquitinase ubiquitin-specific protein 19 (USP19) is known to be involved in deubiquitinating Beclin1, TRAF3, and TRIF for downregulation of the type I IFN signaling. Here, we show that SIAH1, a cellular E3 ubiquitin ligase that is involved in multicellular pathway, is a potent positive regulator of virus-mediated type I IFN signaling that maintains homeostasis within the antiviral immune response by targeting USP19. In the early stages of virus infection, stabilized SIAH1 directly interacts with the USP19 and simultaneously mediates K27-linked ubiquitination of 489, 490, and 610 residues of USP19 for proteasomal degradation. Additionally, we found that USP19 specifically interacts with MAVS and deubiquitinates K63-linked ubiquitinated MAVS for negative regulation of type I IFN signaling. Ultimately, we identified that SIAH1-mediated degradation of USP19 reversed USP19-mediated deubiquitination of MAVS, Beclin1, TRAF3, and TRIF, resulting in the activation of antiviral immune responses. Taken together, these findings provide new insights into the molecular mechanism of USP19 and SIAH1, and suggest a critical role of SIAH1 in antiviral immune response and homeostasis.

CA-CAS-01-A: A Permissive Cell Line for Isolation and Live Attenuated Vaccine Development Against African Swine Fever Virus.

African swine fever virus (ASFV) is the causative agent of the highly lethal African swine fever disease that affects domestic pigs and wild boars. In spite of the rapid spread of the virus worldwide, there is no licensed vaccine available. The lack of a suitable cell line for ASFV propagation hinders the development of a safe and effective vaccine. For ASFV propagation, primary swine macrophages and monocytes have been widely studied. However, obtaining these cells can be time-consuming and expensive, making them unsuitable for mass vaccine production. The goal of this study was to validate the suitability of novel CA-CAS-01-A (CAS-01) cells, which was identified as a highly permissive cell clone for ASFV replication in the MA-104 parental cell line for live attenuated vaccine development. Through a screening experiment, maximum ASFV replication was observed in the CAS-01 cell compared to other sub-clones of MA-104 with 14.89 and log10 7.5 ± 0.15 Ct value and TCID50/ml value respectively. When CAS-01 cells are inoculated with ASFV, replication of ASFV was confirmed by Ct value for ASFV DNA, HAD50/ml assay, TCID50/ml assay, and cytopathic effects and hemadsoption were observed similar to those in primary porcine alveolar macrophages after 5th passage. Additionally, we demonstrated stable replication and adaptation of ASFV over the serial passage. These results suggest that CAS-01 cells will be a valuable and promising cell line for ASFV isolation, replication, and development of live attenuated vaccines.

D-galacto-D-mannan-mediated Dectin-2 activation orchestrates potent cellular and humoral immunity as a viral vaccine adjuvant.

Introduction: Conventional foot-and-mouth disease (FMD) vaccines have been developed to enhance their effectiveness; however, several drawbacks remain, such as slow induction of antibody titers, short-lived immune response, and local side effects at the vaccination site. Therefore, we created a novel FMD vaccine that simultaneously induces cellular and humoral immune responses using the Dectin-2 agonist, D-galacto-D-mannan, as an adjuvant. Methods: We evaluated the innate and adaptive (cellular and humoral) immune responses elicited by the novel FMD vaccine and elucidated the signaling pathway involved both in vitro and in vivo using mice and pigs, as well as immune cells derived from these animals. Results: D-galacto-D-mannan elicited early, mid-, and long-term immunity via simultaneous induction of cellular and humoral immune responses by promoting the expression of immunoregulatory molecules. D-galacto-D-mannan also enhanced the immune response and coordinated vaccine-mediated immune response by suppressing genes associated with excessive inflammatory responses, such as nuclear factor kappa B, via Sirtuin 1 expression. Conclusion: Our findings elucidated the immunological mechanisms induced by D-galacto-D-mannan, suggesting a background for the robust cellular and humoral immune responses induced by FMD vaccines containing D-galacto-D-mannan. Our study will help to facilitate the improvement of conventional FMD vaccines and the design of next-generation FMD vaccines.

The African Swine Fever Virus Virulence Determinant DP96R Suppresses Type I IFN Production Targeting IRF3.

DP96R of African swine fever virus (ASFV), also known as uridine kinase (UK), encodes a virulence-associated protein. Previous studies have examined DP96R along with other genes in an effort to create live attenuated vaccines. While experiments in pigs have explored the impact of DP96R on the pathogenicity of ASFV, the precise molecular mechanism underlying this phenomenon remains unknown. Here, we describe a novel molecular mechanism by which DP96R suppresses interferon regulator factor-3 (IRF3)-mediated antiviral immune responses. DP96R interacts with a crucial karyopherin (KPNA) binding site within IRF3, disrupting the KPNA-IRF3 interaction and consequently impeding the translocation of IRF3 to the nucleus. Under this mechanistic basis, the ectopic expression of DP96R enhances the replication of DNA and RNA viruses by inhibiting the production of IFNs, whereas DP96R knock-down resulted in higher IFNs and IFN-stimulated gene (ISG) transcription during ASFV infection. Collectively, these findings underscore the pivotal role of DP96R in inhibiting IFN responses and increase our understanding of the relationship between DP96R and the virulence of ASFV.

Medial sesamoid position in moderate to severe hallux valgus: Correlation between three radiographic measurements.

BACKGROUND: In hallux valgus surgery, it is essential to accurately assess the position of the sesamoids both pre- and postoperatively. Weight-bearing foot anteroposterior, tangential sesamoid, and semi-weight-bearing computed tomography axial views are radiographic methods used to assess the medial sesamoid position. This study aimed to measure the medial sesamoid position and evaluate the correlation between these three radiographic methods. METHODS: This retrospective study comprised 59 feet from 49 patients who underwent hallux valgus surgery. The mean age of patients was 54.6 (range, 22-70) years. We took preoperative and postoperative measurements using the weight-bearing anteroposterior, tangential sesamoid, and semi-weight-bearing computed tomography axial views to assess the medial sesamoid position. RESULTS: The mean grades of the medial sesamoid position preoperatively and 6 months postoperatively were 2.5 and 0.8, 1.6 and 0.4, and 1.3 and 0.3 points based on the anteroposterior, tangential sesamoid, and computed tomography axial views, respectively (P < 0.001). Preoperatively, there was a strong positive correlation between the computed tomography axial and tangential sesamoid views (P < 0.001, r = 0.645) and anteroposterior and computed tomography axial views (P < 0.001, r = 0.468). In contrast, the tangential sesamoid and anteroposterior views showed a weak positive correlation (P = 0.03, r = 0.283). Six months postoperatively, there was a positive correlation between the computed tomography axial and tangential sesamoid views (P < 0.001, r = 0.473), anteroposterior and computed tomography axial views (P < 0.001, r = 0.470), and tangential sesamoid and anteroposterior views (P < 0.001, r = 0.480). CONCLUSIONS: We observed that the anteroposterior view exhibited a higher degree of medial sesamoid position displacement than the computed tomography axial and tangential sesamoid views. For the preoperative evaluation of the medial sesamoid position, the correlation between the computed tomography axial and tangential sesamoid views was stronger than that between the tangential sesamoid and anteroposterior views. However, all three views showed strong correlations postoperatively.

Advances in Heterostructures for Optoelectronic Devices: Materials, Properties, Conduction Mechanisms, Device Applications.

Atomically thin 2D transition metal dichalcogenides (TMDs) have recently been spotlighted for next-generation electronic and photoelectric device applications. TMD materials with high carrier mobility have superior electronic properties different from bulk semiconductor materials. 0D quantum dots (QDs) possess the ability to tune their bandgap by composition, diameter, and morphology, which allows for a control of their light absorbance and emission wavelength. However, QDs exhibit a low charge carrier mobility and the presence of surface trap states, making it difficult to apply them to electronic and optoelectronic devices. Accordingly, 0D/2D hybrid structures are considered as functional materials with complementary advantages that may not be realized with a single component. Such advantages allow them to be used as both transport and active layers in next-generation optoelectronic applications such as photodetectors, image sensors, solar cells, and light-emitting diodes. Here, recent discoveries related to multicomponent hybrid materials are highlighted. Research trends in electronic and optoelectronic devices based on hybrid heterogeneous materials are also introduced and the issues to be solved from the perspective of the materials and devices are discussed.

Mucosal Administration of Lactobacillus casei Surface-Displayed HA1 Induces Protective Immune Responses against Avian Influenza A Virus in Mice.

Avian influenza is a serious threat to both public health and the poultry industry worldwide. This respiratory virus can be combated by eliciting robust immune responses at the site of infection through mucosal immunization. Recombinant probiotics, specifically lactic acid bacteria, are safe and effective carriers for mucosal vaccines. In this study, we engineered recombinant fusion protein by fusing the hemagglutinin 1 (HA1) subunit of the A/Aquatic bird/Korea/W81/2005 (H5N2) with the Bacillus subtilis poly γ-glutamic acid synthetase A (pgsA) at the surface of Lactobacillus casei (pgsA-HA1/L. casei). Using subcellular fractionation and flow cytometry we confirmed the surface localization of this fusion protein. Mucosal administration of pgsA-HA1/L. casei in mice resulted in significant levels of HA1-specific serum IgG, mucosal IgA and neutralizing antibodies against the H5N2 virus. Additionally, pgsA-HA1/L. casei-induced systemic and local cell-mediated immune responses specific to HA1, as evidenced by an increased number of IFN-γ and IL-4 secreting cells in the spleens and higher levels of IL-4 in the local lymphocyte supernatants. Finally, mice inoculated with pgsA-HA1/L. casei were protected against a 10LD50 dose of the homologous mouse-adapted H5N2 virus. These results suggest that mucosal immunization with L. casei displaying HA1 on its surface could be a potential strategy for developing a mucosal vaccine against other H5 subtype viruses.

Gadd45ß is critical for regulation of type I interferon signaling by facilitating G3BP-mediated stress granule formation.

Stress granules (SGs) constitute a signaling hub that plays a critical role in type I interferon responses. Here, we report that growth arrest and DNA damage-inducible beta (Gadd45ß) act as a positive regulator of SG-mediated interferon signaling by targeting G3BP upon RNA virus infection. Gadd45ß deficiency markedly impairs SG formation and SG-mediated activation of interferon signaling in vitro. Gadd45ß knockout mice are highly susceptible to RNA virus infection, and their ability to produce interferon and cytokines is severely impaired. Specifically, Gadd45ß interacts with the RNA-binding domain of G3BP, leading to conformational expansion of G3BP1 via dissolution of its autoinhibitory electrostatic intramolecular interaction. The acidic loop 1- and RNA-binding properties of Gadd45ß markedly increase the conformational expansion and RNA-binding affinity of the G3BP1-Gadd45ß complex, thereby promoting assembly of SGs. These findings suggest a role for Gadd45ß as a component and critical regulator of G3BP1-mediated SG formation, which facilitates RLR-mediated interferon signaling.

Anhydrous Alum Inhibits α-MSH-Induced Melanogenesis by Down-Regulating MITF via Dual Modulation of CREB and ERK.

Melanogenesis, the intricate process of melanin synthesis, is central to skin pigmentation and photoprotection and is regulated by various signaling pathways and transcription factors. To develop potential skin-whitening agents, we used B16F1 melanoma cells to investigate the inhibitory effects of anhydrous alum on melanogenesis and its underlying molecular mechanisms. Anhydrous alum (KAl(SO4)2) with high purity (>99%), which is generated through the heat-treatment of hydrated alum (KAl(SO4)2·12H2O) at 400 °C, potentiates a significant reduction in melanin content without cytotoxicity. Anhydrous alum downregulates the master regulator of melanogenesis, microphthalmia-associated transcription factor (MITF), which targets key genes involved in melanogenesis, thereby inhibiting α-melanocyte-stimulating hormone (α-MSH)-induced melanogenesis. Phosphorylation of the cAMP response element-binding protein, which acts as a co-activator of MITF gene expression, is attenuated by anhydrous alum, resulting in compromised MITF transcription. Notably, anhydrous alum promoted extracellular signal-regulated kinase phosphorylation, leading to the impaired nuclear localization of MITF. Overall, these results demonstrated the generation and mode of action of anhydrous alum in B16F1 cells, which constitutes a promising option for cosmetic or therapeutic use.

Prediction of Stent Failure for Malignant Ureteral Obstruction in Non-Urological Cancer.

PURPOSE: To analyze prognostic factors associated with ureteral stent failure and to develop a prediction model for malignant ureteral obstruction (MUO) in patients with non-urological cancers. MATERIALS AND METHODS: We retrospectively reviewed patients with non-urological cancers who underwent ureteral stenting or percutaneous nephrostomy (PCN) for MUO between 2006 and 2014. Variables predicting stent failure were identified using Cox regression analysis. RESULTS: Of the 743 patients, 468 (63.0%) underwent ureteral stenting only, and 275 (37.0%) underwent PCN owing to technical (n=215) or functional (n=60) stent failure. The median overall survival was 4 [interquartile range (IQR) 1-11] months, and the median interval duration to stent failure was 2 (IQR 0-7) months. In univariate analysis, lower gastrointestinal cancer, previous radiotherapy to the pelvis, bladder invasion, lower ureteral obstruction, and low previous estimated glomerular filtration rate (eGFR) (<30 mL/min/1.73 m²) were significantly associated with a decreased survival rate. In multivariate analysis, bladder invasion and previous eGFR were significant predictors. With these two predictors, we divided patients into three groups based on their presence: low-risk (neither factor; n=516), intermediate-risk (one factor; n=206), and high-risk (both factors; n=21). The median stent failure-free survival rates of patients in the low-, intermediate-, and high-risk groups were 26 (8-unreached), 1 (0-18), and 0 (0-0) months, respectively (p<0.001). CONCLUSION: In cases of ureteral obstruction caused by non-urological cancers, patients with bladder invasion and a low eGFR showed poor stent failure-free survival. Therefore, PCN should be considered the primary procedure for these patients.

African swine fever virus B175L inhibits the type I interferon pathway by targeting STING and 2'3'-cGAMP.

IMPORTANCE: African swine fever virus (ASFV), the only known DNA arbovirus, is the causative agent of African swine fever (ASF), an acutely contagious disease in pigs. ASF has recently become a crisis in the pig industry in recent years, but there are no commercially available vaccines. Studying the immune evasion mechanisms of ASFV proteins is important for the understanding the pathogenesis of ASFV and essential information for the development of an effective live-attenuated ASFV vaccines. Here, we identified ASFV B175L, previously uncharacterized proteins that inhibit type I interferon signaling by targeting STING and 2'3'-cGAMP. The conserved B175L-zf-FCS motif specifically interacted with both cGAMP and the R238 and Y240 amino acids of STING. Consequently, this interaction interferes with the interaction of cGAMP and STING, thereby inhibiting downstream signaling of IFN-mediated antiviral responses. This novel mechanism of B175L opens a new avenue as one of the ASFV virulent genes that can contribute to the advancement of ASFV live-attenuated vaccines.

The novel immunobiotic Clostridium butyricum S-45-5 displays broad-spectrum antiviral activity in vitro and in vivo by inducing immune modulation.

Clostridium butyricum is known as a probiotic butyric acid bacterium that can improve the intestinal environment. In this study, we isolated a new strain of C. butyricum from infant feces and evaluated its physiological characteristics and antiviral efficacy by modulating the innate immune responses in vitro and in vivo. The isolated C. butyricum S-45-5 showed typical characteristics of C. butyricum including bile acid resistance, antibacterial ability, and growth promotion of various lactic acid bacteria. As an antiviral effect, C. butyricum S-45-5 markedly reduced the replication of influenza A virus (PR8), Newcastle Disease Virus (NDV), and Herpes Simplex Virus (HSV) in RAW264.7 cells in vitro. This suppression can be explained by the induction of antiviral state in cells by the induction of antiviral, IFN-related genes and secretion of IFNs and pro-inflammatory cytokines. In vivo, oral administration of C. butyricum S-45-5 exhibited prophylactic effects on BALB/c mice against fatal doses of highly pathogenic mouse-adapted influenza A subtypes (H1N1, H3N2, and H9N2). Before challenge with influenza virus, C. butyricum S-45-5-treated BALB/c mice showed increased levels of IFN-ß, IFN-γ, IL-6, and IL-12 in serum, the small intestine, and bronchoalveolar lavage fluid (BALF), which correlated with observed prophylactic effects. Interestingly, after challenge with influenza virus, C. butyricum S-45-5-treated BALB/c mice showed reduced levels of pro-inflammatory cytokines and relatively higher levels of anti-inflammatory cytokines at day 7 post-infection. Taken together, these findings suggest that C. butyricum S-45-5 plays an antiviral role in vitro and in vivo by inducing an antiviral state and affects immune modulation to alleviate local and systemic inflammatory responses caused by influenza virus infection. Our study provides the beneficial effects of the new C. butyricum S-45-5 with antiviral effects as a probiotic.

Serological Conversion through a Second Exposure to Inactivated Foot-and-Mouth Disease Virus Expressing the JC Epitope on the Viral Surface.

Foot-and-mouth disease (FMD) is a fatal contagious viral disease that affects cloven-hoofed animals and causes severe economic damage at the national level. There are seven serotypes of the causative foot-and-mouth disease virus (FMDV), and type O is responsible for serious outbreaks and shows a high incidence. Recently, the Cathay, Southeast Asia (SEA), and ME-SA (Middle East-South Asia) topotypes of type O have been found to frequently occur in Asia. Thus, it is necessary to develop candidate vaccines that afford protection against these three different topotypes. In this study, an experimental FMD vaccine was produced using a recombinant virus (TWN-JC) with the JC epitope (VP1 140-160 sequence of the O/SKR/Jincheon/2014) between amino acid 152 and 153 of VP1 in TWN-R. Immunization with this novel vaccine candidate was found to effectively protect mice against challenge with the three different topotype viruses. Neutralizing antibody titers were considerably higher after a second vaccination. The serological differences between the topotype strains were identified in guinea pigs and swine. In conclusion, a significant serological difference was observed at 56 days post-vaccination between animals that received the TWN-JC vaccine candidate and those that received the positive control virus (TWN-R). The TWN-JC vaccine candidate induced IFNγ and IL-12B.

Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT.

Gastrointestinal stromal tumors (GISTs) frequently show KIT mutations, accompanied by overexpression and aberrant localization of mutant KIT (MT-KIT). As previously established by multiple studies, including ours, we confirmed that MT-KIT initiates downstream signaling in the Golgi complex. Basic leucine zipper nuclear factor 1 (BLZF1) was identified as a novel MT-KIT-binding partner that tethers MT-KIT to the Golgi complex. Sustained activation of activated transcription factor 6 (ATF6), which belongs to the unfolded protein response (UPR) family, alleviates endoplasmic reticulum (ER) stress by upregulating chaperone expression, including heat shock protein 90 (HSP90), which assists in MT-KIT folding. BLZF1 knockdown and ATF6 inhibition suppressed both imatinib-sensitive and -resistant GIST in vitro. ATF6 inhibitors further showed potent antitumor effects in GIST xenografts, and the effect was enhanced with ER stress-inducing drugs. ATF6 activation was frequently observed in 67% of patients with GIST (n = 42), and was significantly associated with poorer relapse-free survival (P = 0.033). Overall, GIST bypasses ER quality control (QC) and ER stress-mediated cell death via UPR activation and uses the QC-free Golgi to initiate signaling.

MARCH5-dependent NLRP3 ubiquitination is required for mitochondrial NLRP3-NEK7 complex formation and NLRP3 inflammasome activation.

The NLRP3 inflammasome plays a key role in responding to pathogens, and endogenous damage and mitochondria are intensively involved in inflammasome activation. The NLRP3 inflammasome forms multiprotein complexes and its sequential assembly is important for its activation. Here, we show that NLRP3 is ubiquitinated by the mitochondria-associated E3 ligase, MARCH5. Myeloid cell-specific March5 conditional knockout (March5 cKO) mice failed to secrete IL-1ß and IL-18 and exhibited an attenuated mortality rate upon LPS or Pseudomonas aeruginosa challenge. Macrophages derived from March5 cKO mice also did not produce IL-1ß and IL-18 after microbial infection. Mechanistically, MARCH5 interacts with the NACHT domain of NLRP3 and promotes K27-linked polyubiquitination on K324 and K430 residues of NLRP3. Ubiquitination-defective NLRP3 mutants on K324 and K430 residues are not able to bind to NEK7, nor form NLRP3 oligomers leading to abortive ASC speck formation and diminished IL-1ß production. Thus, MARCH5-dependent NLRP3 ubiquitination on the mitochondria is required for NLRP3-NEK7 complex formation and NLRP3 oligomerization. We propose that the E3 ligase MARCH5 is a regulator of NLRP3 inflammasome activation on the mitochondria.