Biology subjects

Noerenberg, M.

Publications and source records attributed to Noerenberg, M..

9 recordsLinked to original sources

bioRxiv2025

Pre-assembly of biomolecular condensate seeds drives RSV replication

During infection many RNA viruses, including respiratory syncytial virus (RSV), form specialized biomolecular condensates, inclusion bodies (IBs), where viral transcription and replication occur1-4. Paradoxically, high protein concentrations are typically required for condensate nucleation5, yet attaining sufficient protein levels in infection is thought to require IBs for viral transcription and replication. To uncover how viruses solve this paradox to establish IBs, we visualized early infection of RSV in real-time with single genomic viral ribonucleoprotein (vRNP) resolution. Our results reveal that IBs are nucleated from infecting vRNPs rather than de novo in the cytoplasm. IB nucleation further requires in-virion pre-assembly of viral protein-protein interaction networks on vRNPs to form pre-replication centers (PRCs). PRCs are potent condensate nucleation seeds due to their resistance to disassembly and efficient recruitment of newly-synthesized viral proteins. The high protein affinity of PRCs also results in increased polymerase complex association, allowing efficient viral transcription even in the absence of IBs. Together, these activities create a feed-forward loop that drives rapid IB formation. Intriguingly, PRC assembly depends on in-virion viral protein levels and is highly heterogeneous among virions, explaining cell-to-cell heterogeneity in infection progression, and identifying heterogeneous virions as the origin of infection heterogeneity. Together, our results show that in-virion pre-assembly of PRCs kick-starts viral condensate nucleation upon host-cell entry, and explains cell-to-cell heterogeneity in RSV infection.

microbiology↗

bioRxiv2024

XRN1 supplies free nucleotides to feed alphavirus replication

Several RNA viruses induce widespread degradation of cellular mRNAs upon infection; however, the biological significance and mechanistic details of this phenomenon remain unknown. Here, we make use of a model alphavirus, Sindbis virus (SINV), to fill this knowledge gap. We found that SINV triggers cellular RNA decay through the exonuclease XRN1 and the 5-to-3 degradation machinery (5-3DM). These proteins accumulate at viral replication organelles (VROs) and interact with the non-structural protein 1 (nsP1), bringing mRNA degradation into proximity with vRNA synthesis. Our data suggest that monophosphate nucleotides released by cellular RNA decay are recycled through the salvage pathway to feed viral replications. Our work thus reveals a fundamental connection between cellular mRNA degradation and viral replication via nucleotides repurposing. Research highlightsO_LI5-3 RNA decay is essential for the replication of a wide range of viruses. C_LIO_LIXRN1 directly interacts with transcripts which are degraded during infection. C_LIO_LIRNA decay factors and salvage pathway members localise to viral factories. C_LIO_LISupplying nucleosides to several 5-3DM deficient cells facilitates SINV infection. C_LI

biochemistry↗

bioRxiv2024

Uukuniemi virus infection causes a pervasive remodelling of the RNA-binding proteome in tick cell cultures

Cellular RNA-binding proteins (RBPs) are pivotal for the viral lifecycle, mediating key host-virus interactions that promote or repress virus infection. While these interactions have been largely studied in the vertebrate host, no comprehensive analyses of protein-RNA interactions occurring in cells of arbovirus vectors, in particular ticks, have been performed to date. Here we systematically identified the responses of the RNA-binding proteome (RBPome) to infection with a prototype bunyavirus (Uukuniemi virus; UUKV) in tick cells and discovered changes in RNA-binding activity for 283 proteins. In an orthogonal approach, we analysed the composition of the viral ribonucleoprotein by immunoprecipitation of UUKV nucleocapsid protein (N) in infected cells. We found many tick RBPs that are regulated by UUKV infection and associate with viral nucleocapsid protein complexes. We confirmed experimentally that these RBPs impact UUKV infection. This includes the tick homolog of topoisomerase 3B (TOP3B), a protein able to manipulate the topology of RNA, which showed an effect on viral particle production. Our data thus reveals the first protein-RNA interaction map for infected tick cells. Research highlightsO_LIUUKV RNAs interact with nearly three hundred tick cell RBPs. C_LIO_LIDemonstrated an enrichment of N protein interactors within the upregulated RIC data suggesting a direct involvement in viral RNA metabolism and translation. C_LIO_LIDeveloped a robust methodology to silence gene expression in tick cell cultures. C_LIO_LIThe TOP3B complex facilitates efficient packaging of UUKV virions. C_LI

microbiology↗

bioRxiv2023

Incorporation of genome-bound cellular proteins into HIV-1 particles regulates viral infection

The initial steps of the human immunodeficiency virus 1 (HIV-1) lifecycle are regulated by cellular RNA-binding proteins (RBPs). To understand the scope of these early host-virus interactions, we developed in virion RNA interactome capture (ivRIC), which allowed the comprehensive and systematic profiling of the proteins that interact with the HIV-1 genomic (g)RNA inside viral particles. ivRIC identified 104 cellular RBPs within the encapsidated HIV-1 ribonucleoprotein, many of which are typically found in the cellular nucleus. Notably, these nuclear RBPs interact with the HIV-1 RBP Rev, suggesting that they associate with HIV-1 gRNA during its nuclear life. Functional assays show that ivRBPs are important for HIV-1, including PURA and PURB, which control viral gene expression and infectivity through interaction with critical sequences in the gRNA. Our characterisation of the composition of the encapsidated ribonucleoprotein of HIV-1 uncovers new host-virus interactions that invokes new mechanisms for controlling HIV-1 infection.

molecular biology↗

bioRxiv2021

Compositional analysis of Sindbis virus ribonucleoproteins reveals an extensive co-opting of key nuclear RNA-binding proteins

RNA is a central molecule for RNA viruses, acting as mRNA and genome. However, the interactions that viral (v)RNA establishes with the host cell is only starting to be elucidated. Here, we determine with unprecedented depth the composition of the ribonucleoproteins (RNPs) of the prototypical arthropod-borne Sindbis virus (SINV) using viral RNA interactome capture. We show that SINV RNAs engage with hundreds of cellular proteins and pathways, including a group of nuclear RNA-binding proteins (RBPs) with unknown roles in infection. Combining subcellular fractionation and proteomics with several orthogonal approaches, we demonstrate that these nuclear RBPs are selectively redistributed to the cytoplasm after infection, where they associate with the viral replication organelles. These nuclear RBPs potently supress viral gene expression, with activities spanning viral species and families. Our study provides a comprehensive and systematic analysis of SINV RNP composition, revealing a network of nuclear RBPs with moonlighting antiviral function. Research highlightsO_LISINV RNAs interact with over four hundred cellular RBPs C_LIO_LISINV induces selective cytoplasmic translocation of a subset of nuclear RBPs C_LIO_LIThese nuclear RBPs display potent antiviral effects C_LIO_LIThe SF3B complex binds to SINV RNA and supresses infection in a splicing-independent manner C_LI

microbiology↗

bioRxiv2021

Absolute quantitation of individual SARS-CoV-2 RNA molecules: a new paradigm for infection dynamics and variant differences

Despite an unprecedented global research effort on SARS-CoV-2, early replication events remain poorly understood. Given the clinical importance of emergent viral variants with increased transmission, there is an urgent need to understand the early stages of viral replication and transcription. We used single molecule fluorescence in situ hybridisation (smFISH) to quantify positive sense RNA genomes with 95% detection efficiency, while simultaneously visualising negative sense genomes, sub-genomic RNAs and viral proteins. Our absolute quantification of viral RNAs and replication factories revealed that SARS-CoV-2 genomic RNA is long-lived after entry, suggesting that it avoids degradation by cellular nucleases. Moreover, we observed that SARS-CoV-2 replication is highly variable between cells, with only a small cell population displaying high burden of viral RNA. Unexpectedly, the B.1.1.7 variant, first identified in the UK, exhibits significantly slower replication kinetics than the Victoria strain, suggesting a novel mechanism contributing to its higher transmissibility with important clinical implications. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/450133v2_ufig1.gif" ALT="Figure 1"> View larger version (55K): org.highwire.dtl.DTLVardef@10f7bf1org.highwire.dtl.DTLVardef@192214dorg.highwire.dtl.DTLVardef@c84916org.highwire.dtl.DTLVardef@1366287_HPS_FORMAT_FIGEXP M_FIG C_FIG In briefBy detecting nearly all individual SARS-CoV-2 RNA molecules, we quantified viral replication and defined cell susceptibility to infection. We discovered that a minority of cells show significantly elevated viral RNA levels and observed slower replication kinetics for the Alpha variant relative to the Victoria strain. Highlights O_LISingle molecule quantification of SARS-CoV-2 replication uncovers early infection kinetics C_LIO_LIThere is substantial heterogeneity between cells in rates of SARS-CoV-2 replication C_LIO_LIGenomic RNA is stable and persistent during the initial stages of infection C_LIO_LIB.1.1.7 variant replicates more slowly than the Victoria strain C_LI

microbiology↗

bioRxiv2021

The RNA sensor MDA5 detects SARS-CoV-2 infection

Human cells respond to infection by SARS-CoV-2, the virus that causes COVID-19, by producing cytokines including type I and III interferons (IFNs) and proinflammatory factors such as IL6 and TNF. IFNs can limit SARS-CoV-2 replication but cytokine imbalance contributes to severe COVID-19. We studied how cells detect SARS-CoV-2 infection. We report that the cytosolic RNA sensor MDA5 was required for type I and III IFN induction in the lung cancer cell line Calu-3 upon SARS-CoV-2 infection. Type I and III IFN induction further required MAVS and IRF3. In contrast, induction of IL6 and TNF was independent of the MDA5-MAVS-IRF3 axis in this setting. We further found that SARS-CoV-2 infection inhibited the ability of cells to respond to IFNs. In sum, we identified MDA5 as a cellular sensor for SARS-CoV-2 infection that induced type I and III IFNs.

immunology↗

bioRxiv2020

Global analysis of protein-RNA interactions in SARS-CoV-2 infected cells reveals key regulators of infection

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes COVID-19. SARS-CoV-2 relies on cellular RNA-binding proteins (RBPs) to replicate and spread, although which RBPs control SARS-CoV-2 infection remains largely unknown. Here, we employ a multi-omic approach to identify systematically and comprehensively which cellular and viral RBPs are involved in SARS-CoV-2 infection. We reveal that the cellular RNA-bound proteome is remodelled upon SARS-CoV-2 infection, having widespread effects on RNA metabolic pathways, non-canonical RBPs and antiviral factors. Moreover, we apply a new method to identify the proteins that directly interact with viral RNA, uncovering dozens of cellular RBPs and six viral proteins. Amongst them, several components of the tRNA ligase complex, which we show regulate SARS-CoV-2 infection. Furthermore, we discover that available drugs targeting host RBPs that interact with SARS-CoV-2 RNA inhibit infection. Collectively, our results uncover a new universe of host-virus interactions with potential for new antiviral therapies against COVID-19.

microbiology↗

bioRxiv2018

Understanding RNP remodelling uncovers RBPs functionally required for viral replication

The compendium of RNA-binding proteins (RBPome) has been greatly expanded by the development of RNA-interactome capture (RNA-IC). However, it remains unknown how responsive is the RBPome and whether these responses are biologically relevant. To answer these questions, we created comparative RNA-IC to analyse cells challenged with an RNA virus, called sindbis (SINV). Strikingly, the virus altered the activity of 245 RBPs, many of which were newly discovered by RNA-IC. Mechanistically, alterations in RNA binding upon SINV infection are caused by changes in the subcellular localisation of RBPs and RNA availability. Moreover, RBPome responses are crucial, as perturbation of dynamic RBPs modulates the capacity of the virus to infect the cell. For example, ablation of XRN1 causes cells to be refractory to infection, while GEMIN5 moonlights as a novel antiviral factor. Therefore, RBPome remodelling provides a mechanism by which cells can extensively rewire gene expression in response to physiological cues.\n\nHIGHLIGHTSO_LIA quarter of the RBPome remodels upon SINV infection.\nC_LIO_LIThe remodelling is caused by changes in protein localisation and RNA availability.\nC_LIO_LIRewiring of the RBPome is crucial for viral infection efficacy.\nC_LIO_LIWe discover RBPs with previously unknown anti- or pro-viral activity.\nC_LI

molecular biology↗