Identification of a key glioblastoma candidate gene, FUBP3, based on weighted gene co-expression network analysis.

BACKGROUND: Glioblastoma multiforme (GBM) is the most common aggressive malignant brain tumor. However, the molecular mechanism of glioblastoma formation is still poorly understood. To identify candidate genes that may be connected to glioma growth and development, weighted gene co-expression network analysis (WGCNA) was performed to construct a gene co-expression network between gene sets and clinical characteristics. We also explored the function of the key candidate gene. METHODS: Two GBM datasets were selected from GEO Datasets. The R language was used to identify differentially expressed genes. WGCNA was performed to construct a gene co-expression network in the GEO glioblastoma samples. A custom Venn diagram website was used to find the intersecting genes. The GEPIA website was applied for survival analysis to determine the significant gene, FUBP3. OS, DSS, and PFI analyses, based on the UCSC Cancer Genomics Browser, were performed to verify the significance of FUBP3. Immunohistochemistry was performed to evaluate the expression of FUBP3 in glioblastoma and adjacent normal tissue. KEGG and GO enrichment analyses were used to reveal possible functions of FUBP3. Microenvironment analysis was used to explore the relationship between FUBP3 and immune infiltration. Immunohistochemistry was performed to verify the results of the microenvironment analysis. RESULTS: GSE70231 and GSE108474 were selected from GEO Datasets, then 715 and 694 differentially expressed genes (DEGs) from GSE70231 and GSE108474, respectively, were identified. We then performed weighted gene co-expression network analysis (WGCNA) and identified the most downregulated gene modules of GSE70231 and GSE108474, and 659 and 3915 module genes from GSE70231 and GSE108474, respectively, were selected. Five intersection genes (FUBP3, DAD1, CLIC1, ABR, and DNM1) were calculated by Venn diagram. FUBP3 was then identified as the only significant gene by survival analysis using the GEPIA website. OS, DSS, and PFI analyses verified the significance of FUBP3. Immunohistochemical analysis revealed FUBP3 expression in GBM and adjacent normal tissue. KEGG and GO analyses uncovered the possible function of FUBP3 in GBM. Tumor microenvironment analysis showed that FUBP3 may be connected to immune infiltration, and immunohistochemistry identified a positive correlation between immune cells (CD4 + T cells, CD8 + T cells, and macrophages) and FUBP3. CONCLUSION: FUBP3 is associated with immune surveillance in GBM, indicating that it has a great impact on GBM development and progression. Therefore, interventions involving FUBP3 and its regulatory pathway may be a new approach for GBM treatment.

The effect of DNA repair gene variants on COVID-19 disease: susceptibility, severity, and clinical course.

Oxidative stress (OS), which leads to DNA damage, plays a role in the pathogenesis of Coronavirus disease 2019 (COVID-19). We aimed to evaluate the role of DNA repair gene variants [X-ray repair cross complementing 4 (XRCC4) rs28360071, rs6869366, and X-ray cross-complementary gene 1 (XRCC1) rs25487] in susceptibility to COVID-19 in a Turkish population. We also evaluated its effect on the clinical course of the disease. A total of 300 subjects, including 200 COVID-19 patients and 100 healthy controls, were included in this study. These variants were genotyped using polymerase chain reaction (PCR) and/or PCR-restriction fragment length polymorphism (RFLP) methods. The patients were divided into three groups: those with a mild or severe infection; those who died or lived at the 28-day follow-up; those who required inpatient treatment or intensive care. There were 87 women (43.5%) and 113 men (56.5%) in the patient group. Hypertension was the most common comorbidity (26%). In the patient group, XRCC4 rs6869366 G/G genotype and G allele frequency were increased compared to controls, while XRCC4 rs6869366 G/T and T/T genotype frequencies were found to be higher in controls compared to patients. For XRCC1 rs25487, the A/A and A/G genotypes were significantly associated with COVID-19 disease. All of the patients hospitalized in the intensive care unit had the XRCC4 rs6869366 G/G genotype. In this study, we evaluated for the first time the impact of DNA repair gene variants on COVID-19 susceptibility. Results suggested that XRCC4 rs6869366 and XRCC1 rs25487 were associated with COVID-19 suspectibility and clinical course.

ELF5 is a potential respiratory epithelial cell-specific risk gene for severe COVID-19.

Despite two years of intense global research activity, host genetic factors that predispose to a poorer prognosis of COVID-19 infection remain poorly understood. Here, we prioritise eight robust (e.g., ELF5) or suggestive but unreported (e.g., RAB2A) candidate protein mediators of COVID-19 outcomes by integrating results from the COVID-19 Host Genetics Initiative with population-based plasma proteomics using statistical colocalisation. The transcription factor ELF5 (ELF5) shows robust and directionally consistent associations across different outcome definitions, including a >4-fold higher risk (odds ratio: 4.88; 95%-CI: 2.47-9.63; p-value < 5.0 × 10-6) for severe COVID-19 per 1 s.d. higher genetically predicted plasma ELF5. We show that ELF5 is specifically expressed in epithelial cells of the respiratory system, such as secretory and alveolar type 2 cells, using single-cell RNA sequencing and immunohistochemistry. These cells are also likely targets of SARS-CoV-2 by colocalisation with key host factors, including ACE2 and TMPRSS2. In summary, large-scale human genetic studies together with gene expression at single-cell resolution highlight ELF5 as a risk gene for severe COVID-19, supporting a role of epithelial cells of the respiratory system in the adverse host response to SARS-CoV-2.

FACT subunit SUPT16H associates with BRD4 and contributes to silencing of interferon signaling.

FACT (FAcilitates Chromatin Transcription) is a heterodimeric protein complex composed of SUPT16H and SSRP1, and a histone chaperone participating in chromatin remodeling during gene transcription. FACT complex is profoundly regulated, and contributes to both gene activation and suppression. Here we reported that SUPT16H, a subunit of FACT, is acetylated in both epithelial and natural killer (NK) cells. The histone acetyltransferase TIP60 contributes to the acetylation of SUPT16H middle domain (MD) at lysine 674 (K674). Such acetylation of SUPT16H is recognized by bromodomain protein BRD4, which promotes protein stability of SUPT16H in both epithelial and NK cells. We further demonstrated that SUPT16H-BRD4 associates with histone modification enzymes (HDAC1, EZH2), and further regulates their activation status and/or promoter association as well as affects the relevant histone marks (H3ac, H3K9me3 and H3K27me3). BRD4 is known to profoundly regulate interferon (IFN) signaling, while such function of SUPT16H has never been explored. Surprisingly, our results revealed that SUPT16H genetic knockdown via RNAi or pharmacological inhibition by using its inhibitor, curaxin 137 (CBL0137), results in the induction of IFNs and interferon-stimulated genes (ISGs). Through this mechanism, depletion or inhibition of SUPT16H is shown to efficiently inhibit infection of multiple viruses, including Zika, influenza, and SARS-CoV-2. Furthermore, we demonstrated that depletion or inhibition of SUPT16H also causes the remarkable activation of IFN signaling in NK cells, which promotes the NK-mediated killing of virus-infected cells in a co-culture system using human primary NK cells. Overall, our studies unraveled the previously un-appreciated role of FACT complex in coordinating with BRD4 and regulating IFN signaling in both epithelial and NK cells, and also proposed the novel application of the FACT inhibitor CBL0137 to treat viral infections.

Exploring the potential mechanism of emetine against coronavirus disease 2019 combined with lung adenocarcinoma: bioinformatics and molecular simulation analyses.

BACKGROUND: Patients with lung adenocarcinoma (LUAD) may be more predisposed to coronavirus disease 2019 (COVID-19) and have a poorer prognosis. Currently, there is still a lack of effective anti-LUAD/COVID-19 drugs. Thus, this study aimed to screen for an effective anti-LUAD/COVID-19 drug and explore the potential mechanisms. METHODS: Firstly, we performed differentially expressed gene (DEG) analysis on LUAD transcriptome profiling data in The Cancer Genome Atlas (TCGA), where intersections with COVID-19-related genes were screened out. Then, we conducted Cox proportional hazards analyses on these LUAD/COVID-19 DEGs to construct a risk score. Next, LUAD/COVID-19 DEGs were uploaded on Connectivity Map to obtain drugs for anti-LUAD/COVID-19. Finally, we used network pharmacology, molecular docking, and molecular dynamics (MD) simulation to explore the drug's therapeutic targets and potential mechanisms for anti-LUAD/COVID-19. RESULTS: We identified 230 LUAD/COVID-19 DEGs and constructed a risk score containing 7 genes (BTK, CCL20, FURIN, LDHA, TRPA1, ZIC5, and SDK1) that could classify LUAD patients into two risk groups. Then, we screened emetine as an effective drug for anti-LUAD/COVID-19. Network pharmacology analyses identified 6 potential targets (IL6, DPP4, MIF, PRF1, SERPING1, and SLC6A4) for emetine in anti-LUAD/COVID-19. Molecular docking and MD simulation analyses showed that emetine exhibited excellent binding capacities to DDP4 and the main protease (Mpro) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). CONCLUSIONS: This study found that emetine may inhibit the entry and replication of SARS-CoV-2 and enhance tumor immunity by bounding to DDP4 and Mpro.

Autism genes converge on asynchronous development of shared neuron classes.

Genetic risk for autism spectrum disorder (ASD) is associated with hundreds of genes spanning a wide range of biological functions1-6. The alterations in the human brain resulting from mutations in these genes remain unclear. Furthermore, their phenotypic manifestation varies across individuals7,8. Here we used organoid models of the human cerebral cortex to identify cell-type-specific developmental abnormalities that result from haploinsufficiency in three ASD risk genes-SUV420H1 (also known as KMT5B), ARID1B and CHD8-in multiple cell lines from different donors, using single-cell RNA-sequencing (scRNA-seq) analysis of more than 745,000 cells and proteomic analysis of individual organoids, to identify phenotypic convergence. Each of the three mutations confers asynchronous development of two main cortical neuronal lineages-γ-aminobutyric-acid-releasing (GABAergic) neurons and deep-layer excitatory projection neurons-but acts through largely distinct molecular pathways. Although these phenotypes are consistent across cell lines, their expressivity is influenced by the individual genomic context, in a manner that is dependent on both the risk gene and the developmental defect. Calcium imaging in intact organoids shows that these early-stage developmental changes are followed by abnormal circuit activity. This research uncovers cell-type-specific neurodevelopmental abnormalities that are shared across ASD risk genes and are finely modulated by human genomic context, finding convergence in the neurobiological basis of how different risk genes contribute to ASD pathology.

A host transcriptomic signature for identification of respiratory viral infections in the community.

BACKGROUND: Fever-7 is a test evaluating host mRNA expression levels of IFI27, JUP, LAX, HK3, TNIP1, GPAA1 and CTSB in blood able to detect viral infections. This test has been validated mostly in hospital settings. Here we have evaluated Fever-7 to identify the presence of respiratory viral infections in a Community Health Center. METHODS: A prospective study was conducted in the "Servicio de Urgencias de Atención Primaria" in Salamanca, Spain. Patients with clinical signs of respiratory infection and at least one point in the National Early Warning Score were recruited. Fever-7 mRNAs were profiled on a Nanostring nCounter® SPRINT instrument from blood collected upon patient enrolment. Viral diagnosis was performed on nasopharyngeal aspirates (NPAs) using the Biofire-RP2 panel. RESULTS: A respiratory virus was detected in the NPAs of 66 of the 100 patients enrolled. Median National Early Warning Score was 7 in the group with no virus detected and 6.5 in the group with a respiratory viral infection (P > .05). The Fever-7 score yielded an overall AUC of 0.81 to predict a positive viral syndromic test. The optimal operating point for the Fever-7 score yielded a sensitivity of 82% with a specificity of 71%. Multivariate analysis showed that Fever-7 was a robust marker of viral infection independently of age, sex, major comorbidities and disease severity at presentation (OR [CI95%], 3.73 [2.14-6.51], P < .001). CONCLUSIONS: Fever-7 is a promising host immune mRNA signature for the early identification of a respiratory viral infection in the community.

SARS-COV-2 INFECTION AND LONGITUDINAL FECAL SCREENING IN MALAYAN TIGERS (PANTHERA TIGRIS JACKSONI), AMUR TIGERS (PANTHERA TIGRIS ALTAICA ), AND AFRICAN LIONS (PANTHERA LEO KRUGERI) AT THE BRONX ZOO, NEW YORK, USA.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) emerged as the cause of a global pandemic in 2019-2020. In March 2020, New York City became the epicenter in the United States for the pandemic. On 27 March 2020, a Malayan tiger (Panthera tigris jacksoni) at the Bronx Zoo in New York City developed a cough and wheezing with subsequent inappetence. Over the next week, an additional Malayan tiger and two Amur tigers (Panthera tigris altaica) in the same building and three lions (Panthera leo krugeri) in a separate building also became ill. The index case was anesthetized for diagnostic workup. Physical examination and bloodwork results were unremarkable. Thoracic radiography and ultrasonography revealed a bronchial pattern with peribronchial cuffing and mild lung consolidation with alveolar-interstitial syndrome, respectively. SARS-CoV-2 RNA was identified by real-time, reverse transcriptase PCR (rRT-PCR) on oropharyngeal and nasal swabs and tracheal wash fluid. Cytologic examination of tracheal wash fluid revealed necrosis, and viral RNA was detected in necrotic cells by in situ hybridization, confirming virus-associated tissue damage. SARS-CoV-2 was isolated from the tracheal wash fluid of the index case, as well as the feces from one Amur tiger and one lion. Fecal viral RNA shedding was confirmed in all seven clinical cases and an asymptomatic Amur tiger. Respiratory signs abated within 1-5 days for most animals, although they persisted intermittently for 16 days in the index case. Fecal RNA shedding persisted for as long as 35 days beyond cessation of respiratory signs. This case series describes the clinical presentation, diagnostic evaluation, and management of tigers and lions infected with SARS-CoV-2 and describes the duration of viral RNA fecal shedding in these cases. This report documents the first known natural transmission of SARS-CoV-2 from humans to nondomestic felids.

Disruption of Adaptive Immunity Enhances Disease in SARS-CoV-2-Infected Syrian Hamsters.

Animal models recapitulating human COVID-19 disease, especially severe disease, are urgently needed to understand pathogenesis and to evaluate candidate vaccines and therapeutics. Here, we develop novel severe-disease animal models for COVID-19 involving disruption of adaptive immunity in Syrian hamsters. Cyclophosphamide (CyP) immunosuppressed or RAG2 knockout (KO) hamsters were exposed to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by the respiratory route. Both the CyP-treated and RAG2 KO hamsters developed clinical signs of disease that were more severe than those in immunocompetent hamsters, notably weight loss, viral loads, and fatality (RAG2 KO only). Disease was prolonged in transiently immunosuppressed hamsters and was uniformly lethal in RAG2 KO hamsters. We evaluated the protective efficacy of a neutralizing monoclonal antibody and found that pretreatment, even in immunosuppressed animals, limited infection. Our results suggest that functional B and/or T cells are not only important for the clearance of SARS-CoV-2 but also play an early role in protection from acute disease.IMPORTANCE Syrian hamsters are in use as a model of disease caused by SARS-CoV-2. Pathology is pronounced in the upper and lower respiratory tract, and disease signs and endpoints include weight loss and viral RNA and/or infectious virus in swabs and organs (e.g., lungs). However, a high dose of virus is needed to produce disease, and the disease resolves rapidly. Here, we demonstrate that immunosuppressed hamsters are susceptible to low doses of virus and develop more severe and prolonged disease. We demonstrate the efficacy of a novel neutralizing monoclonal antibody using the cyclophosphamide transient suppression model. Furthermore, we demonstrate that RAG2 knockout hamsters develop severe/fatal disease when exposed to SARS-CoV-2. These immunosuppressed hamster models provide researchers with new tools for evaluating therapies and vaccines and understanding COVID-19 pathogenesis.

Utility of a mainstreamed genetic testing pathway in breast and ovarian cancer patients during the COVID-19 pandemic.

INTRODUCTION: Mainstreamed genetic testing (MGT) obviates the need for a cancer genetics consultation, since trained oncologists (O) and gynaecologists (G) provide counseling, prescribe testing and deliver results. We report results from our MGT program and emphasize its utility during the COVID-19 lockdown, when cancer genetics clinics had suspended their activity. METHODS: An MGT pathway for breast and ovarian cancer (BC/OC) patients was established in Jan-2018 between the Assistance Publique - Hôpitaux de Paris.Sorbonne Université Cancer Genetics team and the Oncology/Gynecology departments at one teaching and two regional hospitals. Trained O + G evaluated patients with the Manchester Scoring System. A 12-point threshold was recommended for testing. Next-generation sequencing of BRCA1, BRCA2, PALB2, RAD51C and RAD51D was performed. Results were delivered to the patient by O/G. Pathogenic variants (PV) carriers were referred to the genetics clinic. Results are reported for the 2nd-Jan-2018 to 1st-June-2020 period. That includes the eight-week COVID-19 lockdown and three-week de-confinement phase 1. RESULTS: Results were available for 231/234 patients. Twenty-eight (12.1%) carried a PV. Of the 27 patients tested during the COVID-19 period, three carried a PV, two in BRCA1 and one in RAD51C. The clinical impact was immediate for the two BRCA1 BC cases undergoing neo-adjuvant chemotherapy, since double mastectomy and salpingo-oophorectomy will now be performed using two-step strategies. CONCLUSIONS: MGT guaranteed care continuity in BC/OC patients during the critical phases of the COVID-19 pandemic, with immediate implications for PV carriers. More broadly, we report for the first time the successful implementation of MGT in France.

Association of Clonal Hematopoiesis of Indeterminate Potential With Inflammatory Gene Expression in Patients With Severe Degenerative Aortic Valve Stenosis or Chronic Postischemic Heart Failure.

Importance: Cytokine release syndrome is a complication of coronavirus disease 2019. Clinically, advanced age and cardiovascular comorbidities are the most important risk factors. Objective: To determine whether clonal hematopoiesis of indeterminate potential (CHIP), an age-associated condition with excess cardiovascular risk defined as the presence of an expanded, mutated somatic blood cell clone in persons without other hematological abnormalities, may be associated with an inflammatory gene expression sensitizing monocytes to aggravated immune responses. Design, Setting, and Participants: This hypothesis-generating diagnostic study examined a cohort of patients with severe degenerative aortic valve stenosis or chronic postinfarction heart failure, as well as age-matched healthy control participants. Single-cell RNA sequencing and analyses of circulating peripheral monocytes was done between 2017 and 2019 to assess the transcriptome of circulating monocytes. Exposures: Severe degenerative aortic valve stenosis or chronic postinfarction heart failure. Main Outcomes and Measures: CHIP-driver sequence variations in monocytes with a proinflammatory signature of genes involved in cytokine release syndrome. Results: The study included 8 patients with severe degenerative aortic valve stenosis, 6 with chronic postinfarction heart failure, and 3 healthy control participants. Their mean age was 75.7 (range, 54-89) years, and 6 were women. Mean CHIP-driver gene variant allele frequency was 4.2% (range, 2.5%-6.9%) for DNMT3A and 14.3% (range, 2.6%-37.4%) for TET2. Participants with DNMT3A or TET2 CHIP-driver sequence variations displayed increased expression of interleukin 1ß (no CHIP-driver sequence variations, 1.6217 normalized Unique Molecular Identifiers [nUMI]; DNMT3A, 5.3956 nUMI; P < .001; TET2, 10.8216 nUMI; P < .001), the interleukin 6 receptor (no CHIP-driver sequence variations, 0.5386 nUMI; DNMT3A, 0.9162 nUMI; P < .001;TET2, 0.5738 nUMI; P < .001), as well as the NLRP3 inflammasome complex (no CHIP-driver sequence variations, 0.4797 nUMI; DNMT3A, 0.9961 nUMI; P < .001; TET2, 1.2189 nUMI; P < .001), plus upregulation of CD163 (no CHIP-driver sequence variations, 0.5239 nUMI; DNMT3A, 1.4722 nUMI; P < .001; TET2, 1.0684 nUMI; P < .001), a cellular receptor capable of mediating infection, macrophage activation syndrome, and other genes involved in cytokine response syndrome. Gene ontology term analyses of regulated genes revealed that the most significantly upregulated genes encode for leukocyte-activation and interleukin-signaling pathways in monocytes of individuals with DNMT3A (myeloid leukocyte activation: log[Q value], -50.1986; log P value, -54.5177; regulation of cytokine production: log[Q value], -21.0264; log P value, -24.1993; signaling by interleukins: log[Q value], -18.0710: log P value, -21.1597) or TET2 CHIP-driver sequence variations (immune response: log[Q value], -36.3673; log P value, -40.6864; regulation of cytokine production: log[Q value], -13.1733; log P value, -16.3463; signaling by interleukins: log[Q value], -12.6547: log P value, -15.7977). Conclusions and Relevance: Monocytes of individuals who carry CHIP-driver sequence variations and have cardiovascular disease appear to be primed for excessive inflammatory responses. Further studies are warranted to address potential adverse outcomes of coronavirus disease 2019 in patients with CHIP-driver sequence variations.