Biology subjects

Taube, S.

Publications and source records attributed to Taube, S..

5 recordsLinked to original sources

bioRxiv2025

Polyamine depletion inhibits norovirus infection by blocking virus-induced apoptosis

Noroviruses are non-enveloped, positive-sense RNA viruses in the family Caliciviridae. Human noroviruses (HNoVs) are the leading cause of viral gastrointestinal disease. Despite their clinical relevance, therapeutic strategies for norovirus infections remain limited, partly due to challenges in cultivating HNoV and limited understanding of virus-host interactions. Polyamines (PAs) are small polycationic metabolites derived from amino acid metabolisms and are essential for diverse cellular functions and important host factors for many RNA viruses. However, their role in norovirus infection has remained elusive. Here, we demonstrate that PAs are critical for productive infection for both HNoV and murine norovirus (MNV), underscoring a conserved and functionally significant dependency across species. Functional analysis in MNV-infected murine macrophages showed that PA depletion did not significantly impair virus attachment and viral genome replication. However, it resulted in a marked reduction in infectious virus titers following the first replication cycle, coinciding with a loss of virus-induced apoptosis and impaired virion release. Exogenous supplementation with two metabolites of the PAs biosynthetic pathway (spermine or spermidine), or induction of apoptosis using a PI3-kinase inhibitor, restored both viral titers and apoptosis in PA-depleted macrophages. In human intestinal enteroids (HIEs), depletion of PAs completely ablated HNoV infection, which was partially rescued adding exogenous PAs. Collectively, our findings establish PAs as a host dependency factor that facilitates norovirus infection, promoting virus-induced apoptosis and egress in the murine system. Furthermore, these results point to PAs metabolism as a potential therapeutic target for antiviral intervention against HNoV infections. ImportanceHNoV are a major cause of acute viral gastroenteritis worldwide, yet the development of effective antivirals and vaccines has been hampered by the lack of small-animal models and robust in vitro systems to propagate the virus. MNV shares key features with HNoV and serves as a valuable surrogate for studying norovirus biology. In this study, we identify host-derived PAs as essential and conserved metabolites required for productive infection for both HNoV and MNV. PAs exert multifactorial effects on norovirus infection, particularly by facilitating virus-induced apoptosis and promoting virion release in MNV-infected cells. Notably, PA depletion using difluoromethylornithine (DFMO), a licensed and well-tolerated inhibitor of PA synthesis, effectively suppressed norovirus infection. Our findings highlight the potential of repurposing DFMO as a host-targeting antiviral, especially for the treatment of chronic norovirus infections in immunocompromised individuals. This work provides new insight into norovirus-host interactions and proposes a readily translatable therapeutic avenue.

microbiology↗

bioRxiv2024

NMR Reveals the Synergistic Roles of Bivalent Metal Ions in Norovirus Infections

Norovirus is the most common cause of acute gastroenteritis worldwide. Murine noroviruses (MNV) are often used as model systems for human noroviruses (HuNoV). Therefore, it is important to identify common and divisive properties. Here, we compare the interactions of human and murine norovirus P-domains with bivalent metal ions. Binding of bivalent metal ions and bile acids to MNV P-domains have been shown to stabilize a contracted ("resting") as opposed to an extended ("raised") capsid conformation. This conformational change has been linked to infectivity, diarrheagenic potential, and immune escape. Likewise, the interaction of bivalent metal ions with human norovirus capsids results in contraction, suggesting a similar underlying mechanism. We used methyl TROSY NMR experiments to study the thermodynamics and kinetics of metal ion binding to P-domains, revealing a highly synergistic interaction with the bile acid glycochenodeoxycholic acid (GCDCA) for MNV. Neutralization assays support this synergistic behavior. It turns out that bivalent metal ion binding to MNV and HuNoV P-domains differs significantly. Therefore, although the transition between "raised" and "resting" capsid conformations and consequential modulation of infectivity appears to be triggered by bivalent metal ions in murine and human noroviruses, the underlying mechanisms must be different.

biochemistry↗

bioRxiv2022

Human norovirus efficiently replicates in differentiated 3D-human intestinal enteroids

Human norovirus (HNoV) accounts for one fifth of all acute viral gastroenteritis worldwide and an economic burden of [~]$60 billion globally. The lack of treatment options against HNoV is in part due to the lack of cultivation systems. Recently, a model of infection in biopsies-derived human intestinal enteroids (HIE) has been described: 3D-HIE are first dispersed in 2D-monolayers and differentiated prior to infection, resulting in a labor-intensive, time-consuming procedure. Here, we present an alternative protocol for HNoV infection of 3D-HIE. We found that 3D-HIE differentiate as efficiently as 2D-monolayers. In addition, immunofluorescence-based quantification of UEA-1, a lectin that stains the villus brush border, revealed that over 90% of differentiated 3D-HIE spontaneously undergo polarity inversion, allowing for viral infection without the need for microinjection. Infection with HNoV GII.4-positive stool samples attained a fold-increase over inoculum of [~]2 Log10 at 2 days post infection or up to 3.5 Log10 when ruxolitinib, a JAK1/2-inhibitor, was added. Treatment of GII.4-infected 3D-HIE with the polymerase inhibitor 2-C-Methylcytidine (2CMC), other antivirals, or with a HNoV-neutralizing antibody showed a reduction in viral infection, suggesting that 3D-HIE are an excellent platform to test anti-infectives. The host response to HNoV was then investigated by RNA sequencing in infected versus uninfected 3D-HIE, in the presence of ruxolitinib to focus on viral-associated signatures. The analysis revealed upregulated hormones and neurotransmitter signal transduction pathways and downregulated inflammatory pathways upon HNoV infection. Overall, 3D-HIE have proven to be a more robust model to study HNoV infection, screen antivirals and investigate host response to HNoV infection. ImportanceHuman norovirus (HNoV) clinical and socio-economic impact calls for immediate actions in the development of anti-infectives. Physiologically-relevant in vitro models are hence needed to study HNoV biology, tropism and mechanism of viral-associated disease but also as a platform to identify antiviral agents. Biopsy-derived human intestinal enteroids are a biomimetic of the intestine and recently described as a model that supports HNoV infection. The established protocol is time-consuming and labor-intensive. Therefore, we sought to develop a simplified and robust alternative model of infection in 3D enteroids that undergo differentiation and spontaneous polarity inversion. Advantages of this model are the shorter experimental time, better infection yield and spatial integrity of the intestinal epithelium. This model is potentially suitable for the study of pathogens that infect intestinal cells from the apical surface but also for unraveling the interactions between intestinal epithelium and indigenous bacteria of the human microbiome.

microbiology↗

bioRxiv2021

Protein secondary structure affects glycan clustering in native mass spectrometry

Infection with human noroviruses (hNoV) for the vast majority of strains requires attachment of the viral capsid to histo blood group antigens (HBGA). The HBGA binding pocket is formed by dimers of the protruding domain (P dimers) of the capsid protein VP1. Several studies have focused on HBGA binding to P dimers, reporting binding affinities and stoichiometries. However, nuclear magnetic resonance spectroscopy (NMR) and native mass spectrometry (MS) analyses yielded incongruent dissociation constants (KD) for binding of HBGAs to P dimers and, in some cases, disagreed whether glycans bind at all. We hypothesized that glycan clustering during electrospray ionization in native MS critically depends on the physicochemical properties of the protein studied. It follows that the choice of the reference protein is crucial. We analyzed carbohydrate clustering using various P dimers and eight non-glycan binding proteins serving as possible references. Data from native and ion mobility MS indicate that the mass fraction of {beta}-sheet has a strong influence on the degree of glycan clustering. Therefore, the determination of specific glycan binding affinities from native MS must be interpreted cautiously.

biophysics↗

bioRxiv2021

Murine norovirus capsid plasticity - Glycochenodeoxycholic acid stabilizes P domain dimers and triggers escape from antibody recognition

The murine norovirus (MNV) capsid protein is the target for various neutralizing antibodies binding to distal tips of its protruding (P)-domain. The bile acid glycochenodeoxycholic acid (GCDCA), an important co-factor for murine norovirus (MNV) infection, has recently been shown to induce conformational changes in surface-loops and a contraction of the virion. Here, we employ protein NMR experiments using stable isotope labeled MNV P-domains to shed light on underlying molecular mechanisms. We observe two separate sets of NMR resonance signals for P-domain monomers and dimers, permitting analysis of the corresponding exchange kinetics. Unlike human norovirus GII.4 P-dimers, which exhibit a half-life in the range of several days, MNV P-dimers are very short lived with a half-life of about 17 s. Addition of GCDCA shifts the equilibrium towards the dimeric form by tightly binding to the P-dimers. In MNV virions GCDCA-mediated stabilization of the dimeric arrangement of P-domains generates a more ordered state, which in turn may entropically assist capsid contraction. Numerous long-range chemical shift perturbations (CSPs) upon addition of GCDCA reflect allosteric conformational changes as a feature accompanying dimer stabilization. In particular, CSPs indicate rearrangement of the EF loop, a target for various neutralizing antibodies. Indeed, treating MNV virions with GCDCA prior to neutralizing antibody exposure abolishes neutralization. These findings advance our understanding of GCDCA-induced structural changes of MNV capsids and experimentally support an intriguing viral immune escape mechanism relying on GCDCA-triggered conformational changes of the P-dimer. Significance StatementThis study sheds light on the role of glycochenodeoxycholic acid (GCDCA) in promoting murine norovirus (MNV) infection and immune escape. Binding of GCDCA to the dimeric P-domain has been well characterized by crystallography and cryo EM studies, showing that upon GCDCA binding, a 90{degrees} rotation of the P-domain occurs, which results in its collapse onto the underlying shell of the virus. Our NMR experiments now reveal P-dimer stability as a new dimension of plasticity of MNV capsids and suggest that capsid contraction is entropically assisted. Conformational changes as a feature of P-dimer stabilization eliminate recognition by neutralizing antibodies, no longer being able to prevent infection. These findings highlight key differences between human and MNV capsid structures, promote our understanding of MNV infection on a molecular level, and reveal a novel immune escape mechanism.

biochemistry↗