Biology subjects

Biering, S. B.

Publications and source records attributed to Biering, S. B..

11 recordsLinked to original sources

bioRxiv2025

Dengue Virus NS1 Binds Ephrin B1 to Trigger Endothelial Dysfunction

Dengue virus (DENV), a member of the Flaviviridae family, is the most prevalent and medically important mosquito-borne viral pathogen. Infected cells secrete the viral non-structural protein 1 (NS1) into the bloodstream, where it can interact with endothelial cells to induce vascular leak. Host factors involved in NS1-mediated endothelial dysfunction are incompletely understood, leading us to investigate the host interactome of DENV NS1 in endothelial cells via a comparative mass spectrometry approach. We identified ephrin B1 (EFNB1) as a critical host factor in NS1-mediated endothelial barrier dysfunction and show that phosphorylation of EFNB1 is necessary for induction of barrier dysfunction. Further, we map the interface of the EFNB1-NS1 complex through biochemical and computational approaches, and we show that EFNB1-Fc-fusion proteins can act as decoys to block NS1-induced barrier dysfunction in vitro and in vivo. This study provides insights into the mechanism of flavivirus NS1-mediated endothelial barrier dysfunction and new avenues to target vascular leak.

microbiology↗

bioRxiv2025

Therapeutic Potential of Cyclodextrins Targeting Dengue Virus and SARS-CoV-2 Infection and Pathogenesis

Cyclodextrins (CDs) are cyclic oligosaccharides with promising therapeutic applications, including antiviral activity. During viral infections, pathogenesis arises not only from viral replication but also from viral proteins that act as "toxins", disrupting cellular barriers and inducing endothelial dysfunction, a hallmark of severe diseases such as dengue and COVID-19. Dengue virus (DENV) NS1 and SARS-CoV-2 Spike proteins induce endothelial hyperpermeability, contributing to severe complications. Here we explored the potential of a panel of 18 CDs in mitigating endothelial dysfunction caused by these viral proteins and evaluated the CDs antiviral activity in vitro and in vivo. The effect of CDs on endothelial hyperpermeability was assessed using a trans-endothelial electrical resistance assay with human pulmonary microvascular endothelial cells exposed to DENV NS1 and SARS-CoV-2 Spike proteins. Antiviral efficacy of CDs was evaluated in Vero cells infected with DENV2 and Calu-3 cells infected with SARS-CoV-2, and in vivo protection was assessed in a lethal DENV2 mouse model. CDs effectively inhibited DENV NS1-induced endothelial hyperpermeability in vitro, demonstrating their potential to counteract NS1-mediated barrier disruption. In the murine model, CD1 treatment provided partial protection against DENV-induced morbidity and mortality. Further, CDs significantly reduced SARS-CoV-2 infection in vitro and inhibited Spike-induced endothelial dysfunction. These findings indicate that CDs can prevent endothelial hyperpermeability induced by DENV NS1 and SARS-CoV-2 Spike proteins and exhibit antiviral activity against SARS-CoV-2, positioning them as promising candidates for mitigating endothelial complications associated with viral infections. Further research is needed to explore the clinical relevance of CDs and their mechanisms of action.

microbiology↗

bioRxiv2024

Flavivirus NS1-triggered endothelial dysfunction promotes virus dissemination

The Flaviviridae are a family of viruses that include the important arthropod-borne human pathogens dengue virus (DENV), West Nile virus, Zika virus, Japanese encephalitis virus, and yellow fever virus. Flavivirus nonstructural protein 1 (NS1) is essential for virus replication but is also secreted from virus-infected cells. Extracellular NS1 acts as a virulence factor during flavivirus infection in multiple ways, including triggering endothelial dysfunction and vascular leak via interaction with endothelial cells. While the role of NS1 in inducing vascular leak and exacerbating pathogenesis is well appreciated, if and how NS1-triggered endothelial dysfunction promotes virus infection remains obscure. Flaviviruses have a common need to disseminate from circulation into specific tissues where virus-permissive cells reside. Tissue-specific dissemination is associated with disease manifestations of a given flavivirus, but mechanisms dictating virus dissemination are unclear. Here we show that NS1-mediated endothelial dysfunction promotes virus dissemination in vitro and in vivo. In mouse models of DENV infection, we show that anti-NS1 antibodies decrease virus dissemination, while the addition of exogenous NS1 promotes virus dissemination. Using an in vitro system, we show that NS1 promotes virus dissemination in two distinct ways: (1) promoting crossing of barriers and (2) increasing infectivity of target cells in a tissue- and virus-specific manner. The capacity of NS1 to modulate infectivity correlates with a physical association between virions and NS1, suggesting a potential NS1-virion interaction. Taken together, our study indicates that flavivirus NS1 promotes virus dissemination across endothelial barriers, providing an evolutionary basis for virus-triggered vascular leak. Author SummaryThe Flaviviridae contain numerous medically important human pathogens that cause potentially life-threatening infections. Over half of the worlds population is at risk of flavivirus infection, and this number is expected to increase as climate change expands the habitats of the arthropod vectors that transmit these flaviviruses. There are few effective vaccines and no therapeutics approved for prevention or treatment of flavivirus infection, respectively. Given these challenges, understanding how and why flaviviruses cause pathogenesis is critical for identifying targets for therapeutic intervention. The secreted nonstructural protein 1 (NS1) of flaviviruses is a conserved virulence factor that triggers endothelial dysfunction in a tissue-specific manner. It is unknown if this endothelial dysfunction provides any benefit for virus infection. Here we provide evidence that NS1-triggered endothelial dysfunction facilitates virus crossing of endothelial barriers and augments infection of target cells in vitro and promotes virus dissemination in vivo. This study provides an evolutionary explanation for flaviviruses evolving the capacity to trigger barrier dysfunction and highlights NS1 and the pathways governing endothelial dysfunction, as therapeutic targets to prevent flavivirus dissemination.

microbiology↗

bioRxiv2024

Antibodies targeting Crimean-Congo hemorrhagic fever virus GP38 limit vascular leak and viral spread

Crimean-Congo hemorrhagic fever virus (CCHFV) is a priority pathogen transmitted by tick bites, with no vaccines or specific therapeutics approved to date. Severe disease manifestations include hemorrhage, endothelial dysfunction, and multiorgan failure. Infected cells secrete the viral glycoprotein GP38, whose extracellular function is presently unknown. GP38 is considered an important target for vaccine and therapeutic design as GP38-specific antibodies can protect against severe disease in animal models, albeit through a currently unknown mechanism of action. Here, we show that GP38 induces endothelial barrier dysfunction in vitro, and that CCHFV infection, and GP38 alone, can trigger vascular leak in a mouse model. Protective antibodies that recognize specific antigenic sites on GP38, but not a protective neutralizing antibody binding the structural protein Gc, potently inhibit endothelial hyperpermeability in vitro and vascular leak in vivo during CCHFV infection. This work uncovers a function of the secreted viral protein GP38 as a viral toxin in CCHFV pathogenesis and elucidates the mode of action of non-neutralizing GP38-specific antibodies.

microbiology↗

bioRxiv2023

The Inflammasome Pathway is Activated by Dengue Virus Non-structural Protein 1 and is Protective During Dengue Virus Infection

Dengue virus (DENV) is a medically important flavivirus causing an estimated 50-100 million dengue cases annually, some of whom progress to severe disease. DENV non-structural protein 1 (NS1) is secreted from infected cells and has been implicated as a major driver of dengue pathogenesis by inducing endothelial barrier dysfunction. However, less is known about how DENV NS1 interacts with immune cells and what role these interactions play. Here we report that DENV NS1 can trigger activation of inflammasomes, a family of cytosolic innate immune sensors that respond to infectious and noxious stimuli, in mouse and human macrophages. DENV NS1 induces the release of IL-1{beta} in a caspase-1 dependent manner. Additionally, we find that DENV NS1-induced inflammasome activation is independent of the NLRP3, Pyrin, and AIM2 inflammasome pathways, but requires CD14. Intriguingly, DENV NS1-induced inflammasome activation does not induce pyroptosis and rapid cell death; instead, macrophages maintain cellular viability while releasing IL-1{beta}. Lastly, we show that caspase-1/11-deficient, but not NLRP3-deficient, mice are more susceptible to lethal DENV infection. Together, these results indicate that the inflammasome pathway acts as a sensor of DENV NS1 and plays a protective role during infection.

immunology↗

bioRxiv2022

Kinase-independent activity of DYRK1A promotes viral entry of highly pathogenic human coronaviruses

Identifying host genes essential for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has the potential to reveal novel drug targets and further our understanding of coronavirus disease 2019 (COVID-19). We previously performed a genome-wide CRISPR/Cas9 screen to identify pro-viral host factors for highly pathogenic human coronaviruses. Very few host factors were required by diverse coronaviruses across multiple cell types, but DYRK1A was one such exception. Although its role in coronavirus infection was completely unknown, DYRK1A encodes Dual Specificity Tyrosine Phosphorylation Regulated Kinase 1A and regulates cell proliferation, and neuronal development, among other cellular processes. Interestingly, individuals with Down syndrome overexpress DYRK1A 1.5-fold and exhibit 5-10x higher hospitalization and mortality rates from COVID-19 infection. Here, we demonstrate that DYRK1A regulates ACE2 and DPP4 transcription independent of its catalytic kinase function to support SARS-CoV, SARS-CoV-2, and MERS-CoV entry. We show that DYRK1A promotes DNA accessibility at the ACE2 promoter and a putative distal enhancer, facilitating transcription and gene expression. Finally, we validate that the pro-viral activity of DYRK1A is conserved across species using cells of monkey and human origin and an in vivo mouse model. In summary, we report that DYRK1A is a novel regulator of ACE2 and DPP4 expression that may dictate susceptibility to multiple highly pathogenic human coronaviruses. Whether DYRK1A overexpression contributes to heightened COVID-19 severity in individuals with Down syndrome through ACE2 regulation warrants further future investigation.

microbiology↗

bioRxiv2022

Molecular Determinants of Tissue Specificity of Flavivirus Nonstructural Protein 1 Interaction with Endothelial Cells

Members of the mosquito-borne flavivirus genus such as dengue (DENV), West Nile (WNV), and Zika (ZIKV) viruses cause distinct diseases and affect different tissues. We previously found that the secreted flaviviral nonstructural protein 1 (NS1) interacts with endothelial cells and disrupts endothelial barrier function in a tissue-specific manner consistent with the disease tropism of the respective viruses. However, the underlying molecular mechanism of this tissue-specific NS1-endothelial cell interaction is not well understood. To elucidate the distinct role(s) that the domains of NS1 ({beta}-roll, wing, and {beta}-ladder) play in NS1 interactions with endothelial cells, we constructed flavivirus NS1 chimeras that exchanged the wing and {beta}-ladder domains in a pair-wise manner between DENV, WNV, and ZIKV NS1. We found that both the NS1 wing and {beta}-ladder domains conferred NS1 tissue-specific endothelial dysfunction, with the wing conferring cell binding and the {beta}-ladder involved in inducing endothelial hyperpermeability as measured by trans-endothelial electrical resistance assay. To narrow down the amino acids dictating cell binding specificity, we utilized the DENV-WNV NS1 chimera and identified residues 91 to 93 (GDI) of DENV NS1 as a molecular motif determining binding specificity. Further, using an in vivo mouse model of localized leak, we found that the GDI motif of the wing domain was essential for triggering DENV NS1-induced vascular leak in mouse dermis. Taken together, we identify molecular determinants of flavivirus NS1 that confer NS1 binding and vascular leak and highlight the importance of the NS1 wing domain for flavivirus pathogenesis. ImportanceFlavivirus NS1 is secreted into the bloodstream from infected cells during a viral infection. Dengue virus NS1 contributes to severe dengue pathology such as endothelial dysfunction and vascular leak independently of the virus. We have shown that multiple flavivirus NS1 proteins result in endothelial dysfunction in a tissue-specific manner consistent with their respective viral tropism. Here, we aimed to identify the molecular determinants that make some, but not other, flavivirus NS1 proteins bind to select endothelial cells in vitro and cause vascular leak in a mouse model. We identified the wing domain of NS1 as a primary determinant conferring differential endothelial dysfunction and vascular leak and narrowed the contributing amino acid residues to a three-residue motif within the wing domain. The insights from this study pave the way for future studies on the effects of flavivirus NS1 on viral dissemination and pathogenesis and offer potential new avenues for antiviral therapies.

microbiology↗

bioRxiv2021

SARS-CoV-2 Spike triggers barrier dysfunction and vascular leak via integrins and TGF-β signaling

Severe COVID-19 is associated with epithelial and endothelial barrier dysfunction within the lung as well as in distal organs. While it is appreciated that an exaggerated inflammatory response is associated with barrier dysfunction, the triggers of this pathology are unclear. Here, we report that cell-intrinsic interactions between the Spike (S) glycoprotein of SARS-CoV-2 and epithelial/endothelial cells are sufficient to trigger barrier dysfunction in vitro and vascular leak in vivo, independently of viral replication and the ACE2 receptor. We identify an S-triggered transcriptional response associated with extracellular matrix reorganization and TGF-{beta} signaling. Using genetic knockouts and specific inhibitors, we demonstrate that glycosaminoglycans, integrins, and the TGF-{beta} signaling axis are required for S-mediated barrier dysfunction. Our findings suggest that S interactions with barrier cells are a contributing factor to COVID-19 disease severity and offer mechanistic insight into SARS-CoV-2 triggered vascular leak, providing a starting point for development of therapies targeting COVID-19 pathogenesis.

microbiology↗

bioRxiv2021

An intranasal ASO therapeutic targeting SARS-CoV-2

The COVID-19 pandemic is exacting an increasing toll worldwide, with new SARS-CoV-2 variants emerging that exhibit higher infectivity rates and that may partially evade vaccine and antibody immunity1. Rapid deployment of non-invasive therapeutic avenues capable of preventing infection by all SARS-CoV-2 variants could complement current vaccination efforts and help turn the tide on the COVID-19 pandemic2. Here, we describe a novel therapeutic strategy targeting the SARS-CoV-2 RNA using locked nucleic acid antisense oligonucleotides (LNA ASOs). We identified an LNA ASO binding to the 5 leader sequence of SARS-CoV-2 ORF1a/b that disrupts a highly conserved stem-loop structure with nanomolar efficacy in preventing viral replication in human cells. Daily intranasal administration of this LNA ASO in the K18-hACE2 humanized COVID-19 mouse model potently (98-99%) suppressed viral replication in the lungs of infected mice, revealing strong prophylactic and treatment effects. We found that the LNA ASO also represses viral infection in golden Syrian hamsters, and is highly efficacious in countering all SARS-CoV-2 "variants of concern" tested in vitro and in vivo, including B.1.427, B.1.1.7, and B.1.351 variants3. Hence, inhaled LNA ASOs targeting SARS-CoV-2 represents a promising therapeutic approach to reduce transmission of variants partially resistant to vaccines and monoclonal antibodies, and could be deployed intranasally for prophylaxis or via lung delivery by nebulizer to decrease severity of COVID-19 in infected individuals. LNA ASOs are chemically stable and can be flexibly modified to target different viral RNA sequences4, and they may have particular impact in areas where vaccine distribution is a challenge, and could be stockpiled for future coronavirus pandemics.

microbiology↗

bioRxiv2021

Genome-wide, bidirectional CRISPR screens identify mucins as critical host factors modulating SARS-CoV-2 infection

SARS-CoV-2 can cause a range of symptoms in infected individuals, from mild respiratory illness to acute respiratory distress syndrome. A systematic understanding of the host factors mediating viral infection or restriction is critical to elucidate SARS-CoV-2 host-pathogen interactions and the progression of COVID-19. To this end, we conducted genome-wide CRISPR knockout and activation screens in human lung epithelial cells with endogenous expression of the SARS-CoV-2 entry factors ACE2 and TMPRSS2. These screens uncovered proviral and antiviral host factors across highly interconnected host pathways, including components implicated in clathrin transport, inflammatory signaling, cell cycle regulation, and transcriptional and epigenetic regulation. We further identified mucins, a family of high-molecular weight glycoproteins, as a prominent viral restriction network. We demonstrate that multiple membrane-anchored mucins are critical inhibitors of SARS-CoV-2 entry and are upregulated in response to viral infection. This functional landscape of SARS-CoV-2 host factors provides a physiologically relevant starting point for new host-directed therapeutics and suggests interactions between SARS-CoV-2 and airway mucins of COVID-19 patients as a host defense mechanism.

genetics↗

bioRxiv2020

Screening a library of FDA-approved and bioactive compounds for antiviral activity against SARS-CoV-2

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has emerged as a major global health threat. The COVID-19 pandemic has resulted in over 80 million cases and 1.7 million deaths to date while the number of cases continues to rise. With limited therapeutic options, the identification of safe and effective therapeutics is urgently needed. The repurposing of known clinical compounds holds the potential for rapid identification of drugs effective against SARS-CoV-2. Here we utilized a library of FDA-approved and well-studied preclinical and clinical compounds to screen for antivirals against SARS-CoV-2 in human pulmonary epithelial cells. We identified 13 compounds that exhibit potent antiviral activity across multiple orthogonal assays. Hits include known antivirals, compounds with anti-inflammatory activity, and compounds targeting host pathways such as kinases and proteases critical for SARS-CoV-2 replication. We identified seven compounds not previously reported to have activity against SARS-CoV-2, including B02, a human RAD51 inhibitor. We further demonstrated that B02 exhibits synergy with remdesivir, the only antiviral approved by the FDA to treat COVID-19, highlighting the potential for combination therapy. Taken together, our comparative compound screening strategy highlights the potential of drug repurposing screens to identify novel starting points for development of effective antiviral mono- or combination therapies to treat COVID-19.

microbiology↗