Virologica Sinica

African swine fever (ASF) is a highly pathogenic disease caused by the African swine fever virus (ASFV) infection, which can affect pigs of all ages and breeds, posing significant threat to the global pig farming industry. The ASFV p30 protein is an early-expressed viral structural protein; however, its function is not fully understood. In this study, the interaction of viral p30 with host TRIM21 was identified. The ectopic TRIM21 inhibited ASFV replication, while knockdown or knockout of TRIM21 promoted ASFV replication. Further, p30 was found to interact with RIG-I-like receptor (RLR) signaling adaptor MAVS, and during ASFV infection, p30-TRIM21-MAVS interacted with each other. Mechanistically, TRIM21 activated the K27 polyubiquitination of MAVS to induce IRF3 mediated type I interferon (IFN) production, whereas p30 counteracted TRIM21 activated MAVS K27 polyubiquitination to evade RLR signaling mediated antiviral IFN induction. In summary, our study revealed a novel function of ASFV p30, and provided new insights into the immune evasion of ASFV.

Background: Human papillomaviruses (HPVs) pose a severe threat to global public health by driving nonmelanoma skin cancer (NMSC) and cervical cancer, with NMSC being one of the most common cancers worldwide. Epidermodysplasia verruciformis (EV) is an inborn error of immunity characterized by an increased susceptibility to persistent infection of cutaneous HPV and a high risk of NMSC. The genetic basis remains unknown in many patients with EV. Methods: We collected four unrelated pedigrees with EV. Genetic analysis identified five variants in JAK1 encoding the Janus kinase 1. Ex vivo models and patient-derived tissue were employed to evaluate the functional effects of JAK1 variants and delineate the pathogenic mechanisms. Results: We identified different variants in JAK1 in four pedigrees with dominant EV. Genetic analysis revealed five novel variants in JAK1, three of which resulted in nonsense-mediated mRNA decay (NMD). Functional assays identified a decreased phosphorylation of the signal transducers and activators of transcription (STATs), impaired interferon responses, and defective T cell activation. Immune dysregulation in patients, characterized by a reduced CD4/CD8 T cell ratio, decreased CD8 naive T cell proportion, and accumulated memory T cells, implies impaired antiviral immunity against HPV. Conclusions: Our findings confirm that JAK1 loss-of-function (LOF) variants underlie susceptibility to cutaneous HPV infection. [Funded by the National Natural Science Foundation of China (81788101, 81230015, 82394420, and 82394423), the National Key Research and Development Program of China (2022YFC2703900), the CAMS Innovation Fund for Medical Sciences (2021-I2M-1-018), and the Regione Lombardia, Italy (Innovative Research Project 1137-2010)].

Dengue virus (DENV) is a major burden to global public health, affecting hundreds of millions annually. Children represent the major proportion of global dengue cases, ranging from asymptomatic or subclinical presentation to dengue fever (DF) and severe dengue hemorrhagic fever or shock syndrome (DHF/DSS). The factors that distinguish this range of disease severity are still poorly understood. To identify biomarkers of severity, we analyzed the plasma proteome of acute DENV infected children including both subclinical and hospitalized cases. Proteins associated with the acute-phase response, innate immune and lysosomal activation, and components of the coagulation cascade showed marked differences between hospitalized and subclinical cases during early infection. Longitudinal profiling demonstrated that endothelial dysfunction emerges early, with PTX3 showing the strongest and most rapid upregulation in hospitalized patients, supporting its potential role as a marker of imminent vascular involvement. When comparing severe (DHF/DSS) and classical DF hospitalized cases, CLEC11A displayed the highest fold change at hospital admittance. We used machine-learning analysis to predict disease severity at the acute phase of infection, distinguishing subclinical from hospitalized cases and patients that develop classical dengue fever or severe disease based on the identified complement regulators and inflammatory markers. The panel of identified plasma proteins shed light on the mechanisms of dengue related disease progression and may provide a handle to predict disease severity based on blood markers present during the acute phase of infection.

MicroRNAs (miRNAs) play essential roles in the posttranscriptional regulation of gene expression in organisms. In the process of synthesizing mature miRNAs from miRNA precursors, the miRNA precursors are cleaved via Dicer at their loop structure, after which the miRNA precursors become mature and regulate transcription. However, the consequences of altering the loop sequence are not fully understood. The silkworm Bombyx mori is a lepidopteran insect with many genetic strains. We identified a mutant of the miRNA miR-3260 whose the part of the loop structure was lacking in a silkworm strain with translucent larval skin. Here, we aimed to analyze the role of wild-type miR-3260 and the influence of the mutation of the loop structure in B. mori. First, we identified the genomic region responsible for the translucent larval skin phenotype and determined that the mutated miR-3260 nucleotide sequences. Then, we predicted the binding partners of wild-type miR-3260 using the RNA hybrid tool and found two juvenile hormone (JH)-related genes as targets of wild-type miR-3260. Next, we assessed the relationships between miR-3260 and JH and found that miR-3260 was highly expressed in the Corpora allata and its expression responded to JH treatment. Meanwhile, miR-3260 mimic and inhibitor did not induce the typical phenotypes associated with JH in B. mori. Then, we compared the dicing products from wild-type and mutant miR-3260 precursors and observed that neither form underwent Dicer-mediated cleavage when the loop structure was altered. These results suggest that loop mutations in the miR-3260 precursor may not influence dicing activity, consistent with the lack of observable phenotypic effects.

IntroductionSevere dengue infection is characterized by endothelial injury and systemic inflammatory complications. To better understand the mechanisms underlying disease severity, we investigated a broad panel of circulating inflammatory and endothelial mediators in patients with clinical dengue infection. MethodsA prospective cross-sectional case-control study was carried out involving 111 dengue patients and 42 healthy controls. Among the dengue cases, 85 were identified as primary, while 26 were classified as secondary dengue infections. Serum levels of endothelial markers (Ang-2, CXCL10, MCP1, TRAIL), acute-phase and liver dysfunction and acute-phase markers (CRP, galectin 3, and serum amyloid protein), systemic inflammatory mediators (MIF, TNF-, IL-1{beta}), mast cell-derived proteases (chymase, tryptase), and tissue repair markers HGF, IL-10, IL-1Ra) were quantified using ELISA and Luminex multiplex assays. Correlations among serum analytes, severity indicators, and haematological markers were also explored ResultsSeveral biomarkers, Ang-2, CXCL10, TRAIL, CRP, MIF, IL-1Ra, TNF-, and chymase showed differential expression across severity groups, indicating coordinated endothelial and inflammatory activation. Stratification of patients with primary-secondary dengue also followed a similar pattern except IL-1{beta}, which had significant differential expression across the cohorts. Ang-2 showed strong positive correlations with markers of hepatic dysfunction, including ALT, AST, and bilirubin, suggesting a link between endothelial injury and liver involvement. ConclusionsSevere dengue is driven by the coordinated activation of endothelial dysfunction, acute-phase responses, mast cell mediators, and counter-regulatory pathways. These processes collectively contribute to vascular leakage and organ injury, reinforcing the value of biomarkers such as Ang-2, CXCL10, CRP, and chymase for severity assessment.

The yerba mate psyllid (Gyropsylla spegazziniana) poses a significant threat to yerba mate crops, causing extensive economic losses. While some ecological aspects as well as control strategies have been studied, its associated viral diversity remains unexplored. Here, by generating the first RNA high-throughput analysis (HTS) of this pest, we explored the G. spegazziniana virome, revealing novel and diverse RNA viruses. We characterized five new viral members belonging to distinct families, with evolutionary cues of beny-like viruses (Benyviridae), picorna-like viruses (Picornaviridae), and sobemo-like viruses (Solemoviridae); which were tentatively named Gyropsylla spegazziniana beny-like virus 1 (GSBlV1), Gyropsylla spegazziniana picorna-like virus 1 (GSPlV1), and Gyropsylla spegazziniana sobemo-like virus 1-3 (GSSlV1-3), respectively. Phylogenetic analysis of the bi-segmented and highly divergent sobemo-like viruses showed a distinctive evolutionary trajectory of its encoding proteins at the periphery of recently reported invertebrate Sobelivirales. The beny-like virus belonged to a cluster of insect-associated beny-like viruse; while the picorna-like virus clustered together with psyllid-associated picorna-like viruses. These results highlight the existence of a complex virome within G. spegazziniana and establish the basis for future studies investigating the ecological roles, evolutionary dynamics, and potential biocontrol applications of these viruses in the G. spegazziniana -yerba mate eco-systems.

PurposeAntibodies to Epstein-Barr virus (EBV) proteins can predict nasopharyngeal carcinoma (NPC) risk. We previously defined a prototype EBNA1 protein panel and multiplex immunoblot assay that distinguishes NPC risk several years pre-diagnosis. Assay throughput and specificity are critical to effectively implement a population-level screening program. Here, we developed a strip test assay - EBNA1 SeroStrip-HT - with an objective to increase throughput and maximize specificity. Experimental DesignEBNA1 full-length (FL) and glycine-alanine repeat deletion mutants (dGAr) were purified from insect and mammalian cells to screen serum IgA/IgG from prospective cohorts in Singapore and Shanghai, China, with known time intervals to NPC diagnosis. Twenty pre-diagnostic sera within 4 years to diagnosis were compared to 96 healthy controls using a nested case-control study design. ResultsIgA to mammalian-derived EBNA1 dGAr achieved 85.0% sensitivity and 94.8% specificity (AUC, 0.939) for NPC status. IgA to insect-derived EBNA1 dGAr showed the same sensitivity (85.0%) and similar specificity (93.8%) (AUC, 0.941). IgA to insect-derived EBNA1 FL had a higher 90% sensitivity, but lower 91.7% specificity (AUC, 0.940). Combining EBNA1 FL and dGAr results showed that subjects positive for both proteins had a 243.67 odds ratio for NPC incidence compared to double-negative scores. ConclusionThis study demonstrated the efficacy of EBNA1 SeroStrip-HT for NPC risk assessment and stratification in high- and intermediate-risk populations, yielding high accuracy and a 12-fold increased throughput over the prototype. The insect system was appropriate for large-scale production of purified EBNA1. Larger, geographically diverse cohorts are warranted to confirm these results, especially in low-incidence populations.

Despite of the highly potent antiretroviral therapies, HIV-1 establishes persistent infection and causes chronic inflammation in AIDS patients. Beyond CD4+ T cells, HIV-1 infects myeloid cells, including circulating monocytes and tissue-resident macrophages, and integrates with host genomes to form stable viral reservoirs. To achieve a functional HIV cure, latency-promoting agents (LPAs) have been developed for the "block-and-lock" strategy to reinforce deep HIV-1 latency and permanently silence proviruses. However, most LPAs have been tested mainly in CD4+ T cells, and their efficacy in myeloid cells remains unclear. In this study, we reported that levosimendan (LSM), a drug approved for clinic use to treat heart failures, is able to inhibit HIV lytic infection and reactivation in myeloid cells. LSM blocked viral lytic reactivation in HIV-1 latently infected monocytic cells (TH89GFP, U1) and microglial cells (HC69). LSM also inhibited HIV infection in human induced pluripotent stem cell (iPSC) derived microglia (iMG), primary human resident liver macrophages (Kupffer cells) as well as human monocyte-derived macrophages (MDMs). Furthermore, we demonstrated that overexpression of a predicted drug target of LSM, the conserved serine/threonine kinase RIOK1 (RIO kinase 1), overcomes LSMs anti-HIV effect. Overall, our studies concluded that LSM is a promising LPA to inhibit HIV-1 infection in myeloid cells in the RIOK1-dependent manner.

The primary route of SARS-CoV-2 entry is via respiratory epithelium. However, many COVID-19 patients developed dermatological lesions, and SARS-CoV-2 RNA has been detected in the patients skin. Inflammatory skin diseases, psoriasis and atopic dermatitis (AD), significantly increased the risk of COVID-19. To evaluate the potential role of skin in SARS-CoV-2 host interactions, we utilized 3D human skin organoids (HSO) generated from human epidermal keratinocytes, as well as neonatal skin explants. HSO were treated with cytokines involved in acute and chronic skin inflammation and cytokine storm in severe COVID-19 disease, TNF-, IL-6, IL-1{beta}, and IFN-{gamma}, individually and in combination. HSO were also treated with Th1 (TNF- + IL-17) and Th2 (IL-4 + IL-13) cocktails inducing pro-psoriasis and pro-AD HSO changes, respectively. All individual cytokines, and especially their combinations, elevated the expression of ACE2 and TMPRSS2 at mRNA/protein levels. The Th2 induced only TMPRSS2, the Th1 predominantly induced ACE2. Topically applied Spike-pseudotyped lentiviral Tomato reporter, which binds ACE2 similarly to SARS-CoV-2, successfully infected control and cytokine-treated HSO as well as neonatal skin explants. Cytokine treatment, especially TNF- + IL-6 + IL-1{beta} + IFN-{gamma} and the Th1, significantly increased viral entry. Transcriptomic analysis further revealed partial overlap between gene expression signatures induced by Spike-mediated entry in inflamed HSO and those observed in lung tissue from COVID-19 patients, supporting the biological relevance of skin models. Together, these findings demonstrate that inflammation enhances the permissiveness of human skin to SARS-CoV-2 entry, suggesting that the skin may represent a previously underappreciated interface in viral host interactions.

Cancer-related genomic instability (GI) may cause genetic alterations in spermatozoa, implying health issues not only in cancer survivors, but also in their children [1, 2]. We therefore studied Loss of Y chromosome (LOY), considered as hallmark of GI [3-15], in spermatozoa and blood from survivors of childhood and testicular cancer (CC, TC), and controls (CTRL). We found that LOY was statistically significantly more frequent in spermatozoa from cancer survivors than in controls (Odds Ratio [OR]=2.2 for CC vs. CTRL and OR=2.4 for TC vs. CTRL). Furthermore, LOY was about an order of magnitude more prevalent in spermatozoa than in blood among 18-53-year-old males within all cohorts. Our findings suggest that LOY in spermatozoa might be a clinically useful marker of GI, reduced fertility and disease predisposition in males. Introducing LOY in spermatozoa as a biomarker opens a new research avenue into disease prevention and the causes and consequences of LOY.

Mpox virus (MPXV) gained increased attention following the declaration of two Public Health Emergencies of International Concern (PHEICs) in 2022 and 2024. The rapid spread of MPXV and the increase in human-to-human transmission highlighted the need for improved diagnostic tools for characterizing infection patterns and transmission dynamics. While PCR is effective for detecting active infections, serological approaches can help identify previous or asymptomatic infections and support retrospective surveillance. However, many serological assays developed during recent outbreaks have not been evaluated in endemic settings such as the Democratic Republic of the Congo (DRC). This study aims to define antigen-specific serological cutoff values to differentiate MPXV-seroreactive individuals from those with other orthopoxvirus (OPXV) exposure or different vaccination histories, specifically for use in the DRC. Here, we analyzed 134 individuals, divided into six distinct cohorts with different exposures. Serum samples were tested using Mesoscale Discovery (MSD) to screen for five MPXV and vaccinia virus (VACV) orthologous antigens: A29L/A27L, A35R/A33R, B6R/B5R, E8L/D8L, and M1R/L1R. Receiver operating characteristic (ROC) analysis identified the best-performing antigens and established seroreactivity cutoff values. A binary composite rule was also evaluated to improve the classification of these results. We identified three MPXV antigens, E8L (cut-off=12.33 AU/mL), A35R (cut-off=5.22 AU/mL), and B6R (cut-off=9.77 AU/mL), that showed the strongest discriminatory performance in the dataset. Collectively, these three antigens form a significant panel that demonstrated clear separation between our mpox survivor cohort and other OPXV-exposed individuals.

Climate change and ecosystem collapse promote geographic expansion of vector-borne diseases, as witnessed by the recent incursions into Spain of the virus responsible for Crimean-Congo hemorrhagic fever (CCHFV). CCHFV is maintained in a tick-vertebrate cycle, principally involving ticks of the genus Hyalomma. Faced with the spread of Hyalomma ticks, and therefore the threat of a natural introduction of CCHFV into Western Europe, appropriate surveillance tools and control measures need to be implemented. It is both within and by the tick that CCHFV is maintained and spread in the environment. Despite prolonged portage of the virus, the tick is not overtly affected by CHFV infection. One of the prerequisites in conceiving control strategies is to understand the molecular mechanisms that intimately link the virus to its arthropod host. Despite the central role of the tick in the biology of CCHFV, these mechanisms are ill-defined, owing in part to the constraints associated with handling CCHFV-infected ticks in biosafety level 4 containment. In this study, we established the network of interactions between the S segment of the RNA genome Hazara virus (HAZV), a BSL-2 model of CCHFV, and Hyalomma proteins using ChIRP-MS technique. We identified 166 tick proteins, 21 of which have been described as RNA-binding proteins. Gene ontology and pathway enrichment analyses revealed that the S segment RNA interacts predominantly with mitochondrial proteins that belong to various mitochondrial metabolic pathways.

Schistosomiasis is a major cause of hepatic fibrosis in endemic regions, yet the host genetic determinants of disease progression remain poorly defined. We aimed to identify genetic drivers and underlying mechanisms of schistosomiasis-induced hepatic fibrosis. We performed a genome-wide association study (GWAS) of 984 Schistosoma japonicum-infected individuals from hyperendemic areas in China followed by multi-omics integration and experimental validation to identify causal genes and fibrogenic pathways. Schistosomiasis-associated fibrosis exhibited a genetic architecture distinct from metabolic and viral liver fibrosis, supporting pathogen-specific mechanisms. Eight novel susceptibility loci were identified, including a genome-wide significant signal at 16p13 (rs73575170, P = 3.9 x 10-8). Integrative mapping linked these loci to 262 genes enriched in liver sinusoidal endothelial cells (P = 5.84 x 10-5) and sphingolipid metabolism pathways (P = 4.19 x 10-5). Notably, Diacylglycerol kinase gamma (DGKG, rs6762330, P = 4.37 x 10-6) emerged as a key candidate, with its expression in peri-granuloma and periportal hepatocytes strongly correlating with fibrosis severity (r = 0.816). In vivo, Dgkg knockout attenuated hepatic fibrosis and immunopathology while restoring cholesterol homeostasis, whereas Dgkg overexpression exacerbated fibrogenesis and increased TNF-{beta} levels tenfold. This study identifies DGKG as a key mediator linking lipid metabolism and immune signaling in schistosomiasis-induced fibrosis, uncovering a pathogen-specific genetic mechanism and providing a potential therapeutic target for infection-associated liver fibrosis.

Alzheimer's disease (AD) is a major cause of dementia, with polygenic risk scores (PRSs) widely used to capture cumulative genetic risk. While PRSs have been associated with cognitive decline, their relevance to clinically accessible measures in general populations is not yet fully established, particularly in non-European cohorts. In this study, we investigated the association between AD PRSs and cognitive function assessed by the Mini-Mental State Examination (MMSE) in a community-dwelling Japanese older population (N = 1,301). Three PRSs were constructed using genome-wide association study (GWAS) summary statistics derived from European and Japanese populations. Among the PRSs, the score based on Japanese GWAS showed the strongest and most consistent association with MMSE score, whereas those based on European GWAS showed weaker or no associations. Stratification analyses further demonstrated that individuals with higher PRS exhibited lower MMSE scores and a higher prevalence of cognitive impairment. Notably, these associations were attenuated after excluding participants with dementia, suggesting that PRS primarily reflects clinically relevant cognitive decline. No significant associations were observed between PRSs and hippocampal volume in our cohort. These findings highlight the importance of population-specific PRS and suggest its potential utility for stratifying cognitive impairment using simple clinical measures in community-based settings.

Neutralizing monoclonal antibodies (mAbs) are a key component of antiviral therapeutics against SARS-CoV-2; however, the contribution of Fc-mediated effector functions remains underexplored. Here, we compare the antiviral activities of the neutralizing and non-neutralizing mAbs CB6 and CR3022, respectively. The Fc regions of both plant-produced mAbs carried nonfucosylated, non-galactosylated complex glycans (pCB6 and pCR3022), and CR3022 was also produced with mammalian-typical galactosylated, fucosylated glycans (mCR3022). pCR3022 exhibited markedly enhanced antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cell-mediated virus inhibition (ADCVI) compared to mCR3022, indicating a significant impact of Fc glycosylation on antiviral activity despite the lack of neutralization. pCB6 exhibited potent neutralization while further enhancing virus clearance through synergistic Fc effector activity. Our findings suggest that Fc-mediated mechanisms, especially ADCC and ADCVI, can contribute substantially to viral control and may be particularly valuable against immune-evasive variants. These results advance our understanding of the functional roles that non-neutralizing antibodies can play in SARS-CoV-2 infection and highlight the potential of Fc glycoengineering to modulate the antiviral efficacy of both neutralizing and non-neutralizing mAbs.

To understand the molecular and cellular mechanisms beyond B-cell depletion with the anti-CD20 monoclonal antibody ocrelizumab, we used comprehensive muti-modal flow cytometry and functional assays in a prospective longitudinal multiple sclerosis (MS) cohort. Ocrelizumab depleted the vast majority of B cells and showed selective effects on different B cells subsets. Analysis of residual/replenished B cells revealed relative enrichment of regulatory B cells like CD27+CD43+ B1 and CD24hiCD38hi transitional B cells, and reduction of CD27+ memory B cell subsets and CD19+IgD+CD27-naive B cells at early time points (1-3 month) and before subsequent infusions at 4-7 months, 11-14 months, and >18 months. CD20+ T cells and peripheral helper T-cells (Tph) were also reduced. RNA sequencing analysis showed B1 cells have significantly higher expression of LGALS1, KCNN4, ITGB1, and IL2RB. Compared to transitional B cells, B1 cells also displayed significantly higher expression of tissue homing molecules ITGAX (CD11c), S100A4, ITGB1, and CXCR3. IL10 signaling pathway is increased in these B cells. Ex vivo B cell functional assays indicated the residual/replenishing B cells were anergic following ocrelizumab, with increased IL10/TNF and IL10/IL6 ratios under BCR stimulation. Ocrelizumab treatment may create a self-reinforcing regulatory circuit: the reduction of Tph cells could alleviate suppression of regulatory B cells, which subsequently expand and further promote regulatory T cell networks via IL2RB, LGALS1, and an increased IL-10 signaling pathway.

Background: The global burden of dengue infection has rising, yet limited data exists on its impact in the Caribbean. We describe the incidence and associates of acute kidney injury in adults and children with dengue at a teaching hospital in Jamaica. Methods: A single-centre retrospective cohort study of admissions with laboratory confirmed dengue infection at University Hospital of the West Indies, Mona Jamaica between January 2023 to November 2024. AKI was defined using Kidney Disease Improving Global Outcomes definitions. Patients were included if aged >1year and had at least 2 creatinine values. Clinical, demographic and laboratory data were abstracted by chart review. Summary statistics were used to describe continuous and categorical data, and logistic regression to determine AKI associations. Stratified analysis was performed by age-group (adults-aged [&ge;] 16, and paediatric-aged <16 years). Results: Analyses included 167 persons, 62% (103) were male, mean age was 26.1{+/-}19.5 years. AKI occurred in 25.8%, 65.1% were KDIGO stage 1. AKI incidence was 30.2% and 18.0% among adults and children respectively. There were 3 in-hospital deaths. People with AKI were older 32{+/-}21.4 vs 24 {+/-}18.4 (p=0.021), and had longer duration of stay [6 vs 4 days (p <0.001)]. Male sex [OR 2.09 (95% CI:0.96-4.59), p=0.064], age per year [OR 1.02 (95% CI:1.01-1.04), p=0.015] symptom duration [OR1.11 (CI 0.99-1.24), p = 0.058], admission bilirubin [OR 1.02 (CI: 1.00-1.04), p = 0.022], NLR [OR 1.09 (CI 1.00-1.18), p = 0.037] were associated with AKI. In adults admission potassium was inversely associated with AKI [OR 0.46 (95% CI 0.21-1.01), p 0.056], while in children admission potassium [OR 3.00 (95% CI 0.88-10.6), p 0.088] was associated with AKI. Conclusion: AKI in dengue hospitalizations is higher than most reports at 25.8%. Targeted public health policy on vector control and early symptom recognition may be needed to improve outcomes.

Background and aimsCirculating complement is associated with occurrence of alcohol-associated hepatitis (AH) and is a potential biomarker to distinguish AH from alcohol cirrhosis (AC). Complement contributes to kidney injury, a condition often occurring in patients with alcohol-associated liver disease (ALD). However, little is known regarding complement in cross talk between liver and kidney in ALD. Here we tested the hypothesis that urinary complement would provide potential biomarkers for ALD and insights into mechanisms of liver-kidney crosstalk in the pathogenesis of ALD. MethodsPlasma and urine were collected at admission from patients with sAH, healthy controls (HC), and heavy drinkers without liver disease (HD) (from the multicenter Alcohol Hepatitis Network) and with AC (from the Northern Ohio Alcohol Center). Urine was subjected to unbiased proteomics analysis and plasma complement assessed by multiplex/ELISA assays. 30- and 90-day mortality was tracked in patients with sAH. ResultsAll three complement activation pathways were perturbed in plasma and urine of patients with sAH and AC compared to HC and HD. Components of the lectin and classical pathways in urine were associated with 30- and 90-day mortality in patients with sAH. When 4 complement proteins were combined, they distinguished sAH from AC (AUC 0.78), equivalent to that of MELD (AUC 0.65). There was no correlation between complement in plasma and urine, suggesting an independent impact of sAH on complement in kidney and liver. ConclusionThe urinary proteome revealed complement protein signatures associated with sAH and AC, providing valuable insights into the potential for complement biomarkers and the mechanisms of liver-kidney crosstalk in ALD.

Ubiquitin-like protein 3 (UBL3) is a post-translational modifier that sorts proteins into small extracellular vesicles and regulates the trafficking of disease-associated proteins such as -synuclein. The structural and dynamic features of the UBL domain that underlie these functions, however, remain poorly understood. Here we performed in silico structural dynamics analysis of the UBL3 UBL domain using an NMR structure ensemble combined with anisotropic network modeling (ANM) and perturbation response scanning (PRS). Principal component analysis and residue-wise fluctuation analysis consistently revealed high flexibility in the C-terminal region of UBL3. Comparative ANM analysis across 20 ubiquitin-like proteins (UBLs) further showed that C-terminal flexibility is a conserved yet variable property within the UBL family. PRS analysis demonstrated that residues forming the central -helix of the {beta}-grasp fold exert greater dynamic control over collective motions than {beta}-sheet residues. Notably, UBL3 displayed the highest helix/sheet PRS effectiveness ratio among all UBLs analyzed, highlighting the prominent dynamic contribution of helix residues in this domain. Together, these results provide a structural basis for understanding UBL3-dependent protein interactions and disease-related mechanisms, and suggest that helix-centered dynamic control in the UBL domain may represent a potential target for modulating UBL3 function.

Coat protein (CP) tertiary structures and their capsid organization of spherical viruses are highly conserved within each virus family. While AlphaFold successfully predicts the tertiary structures of individual CPs, their association to form proper quaternary assemblies cannot be easily accomplished. Here, we report a generalized methodology and associated web-based utility (https://foldavirus.org) that combines AlphaFold predictions of CPs with the knowledge on corresponding icosahedral architectures (e.g., T=1, 3, 4...) based on the known structures from the same virus family to generate associated capsids. The resulting assemblies are subjected to Amber energy minimization to relieve any steric clashes at the inter-subunit interfaces. Significantly, the capsid models are validated by calculating robust Mahalanobis distance using the residue annotations categorized as interface, core and surface amino acids with respect to those observed in the experimentally determined analogous structures. Given the amino acid sequence of CP(s), we successfully generated capsids up to T=9 icosahedral symmetry, including those of Picornaviruses that display pseudo-T=3 symmetry comprising different CPs. As the number of currently available CP sequences are 3-4 orders of magnitude larger than the experimentally determined 3D-structures, this approach bridges the huge gap that exists between the corresponding sequence and structure space.