Biology subjects

Werner, A.-D.

Publications and source records attributed to Werner, A.-D..

4 recordsLinked to original sources

bioRxiv2026

Amino acid Mutations at the Interface of Sudan Virus VP40 alter transport and budding

The matrix protein VP40 of orthoebolaviruses coordinates virion release and downregulates viral RNA synthesis through distinct oligomeric states, including dimers, octamers, and filamentous assemblies. To dissect the contributions of two oligomeric interface residues, L117 and W95, in the Sudan virus (SUDV) VP40 (sVP40), we created variants carrying alanine substitutions and assessed their structural and functional properties. sVP40 L117A failed to form dimers and was predominantly monomeric showing increased structural flexibility, reduced thermal stability together with loss of plasma membrane transport, budding activity, and the ability to regulate viral RNA synthesis. VP40 W95A preserved dimerization but also exhibited increased structural flexibility and reduced thermal stability. Functionally, sVP40 W95A more strongly inhibited viral RNA synthesis and markedly enhanced budding. However, in a transcription- and replication-competent virus-like particle (trVLP) assay, trVLPs produced with sVP40 W95A induced substantially reduced reporter activity in target cells, indicating impaired particle infectivity or functionality and suggesting possible defects in minigenome packaging, entry, or early post-entry steps. These results demonstrate that mutations at key oligomerization interfaces exert distinct structural and functional effects and highlight the requirement for precise oligomerization in coordinating sVP40s dual roles in genome regulation and virion release. By defining the contributions of L117 and W95, this study advances mechanistic understanding of sVP40 function and identifies processes that may serve as targets for antiviral intervention. ImportanceSudan virus (SUDV) causes regular outbreaks in Sub-Sahara Africa with unusually high lethality rates. However, in contrast to the more often occurring Zaire ebolavirus (EBOV), no monoclonal antibodies or vaccines are available and SUDV is generally understudied. The matrix protein VP40 is responsible for the downregulation of viral genome replication and transcription as well as budding. Here, we present structural and functional characterization of the SUDV VP40 interface residues L117 and W95 and show that while both amino acids are crucial for VP40s structural integrity, their functional effects are dramatically different ranging from complete abolishment to improving regulatory and budding activities.

microbiology↗

bioRxiv2026

Lethal Sudan virus infection in IFNAR-/- mice reveals hallmarks of a cytokine storm

Sudan virus (SUDV) is a member of the family Filoviridae, which comprises highly pathogenic viruses associated with unusually high case fatality rates. The development of medical countermeasures against filoviruses, including antivirals, vaccines, and therapeutic antibodies, requires preclinical evaluation in suitable animal models. C57BL/6J IFNAR-/- mice, which lack the type I interferon (IFN-/{beta}) receptor, have been reported to be susceptible to filovirus infections, although their impaired innate immune response may represent a potential limitation of the model. Here, we show that IFNAR-/- mice constitute a suitable model for SUDV infection. Following infection, animals developed a clear clinical disease characterized by significant weight loss and pronounced changes in behaviour and appearance. Mice reached the predefined clinical endpoint 3-5 days post infection. Post mortem analysis of terminal samples revealed high viral loads and viral genome copies in all tested organs as well as in serum, indicating widespread systemic dissemination. Importantly, infection was associated with a marked increase in several key chemokines and cytokines linked to systemic inflammation, consistent with the development of a cytokine storm-like response. Together, these findings demonstrate that SUDV infection in IFNAR-/- mice induces systemic viral dissemination and a pronounced inflammatory response, supporting the suitability of this model for investigating filovirus pathogenesis and infection-associated immune dysregulation.

microbiology↗

bioRxiv2025

Side-by-side evaluation of two mouse models for Crimean-Congo Hemorrhagic Fever Virus infection.

Crimean-Congo hemorrhagic fever virus (CCHFV) is the causative agent of a severe hemorrhagic fever in humans, associated with case fatality rates ranging from 10 to 40%. Due to the lack of approved vaccines or specific antiviral treatments, CCHFV is classified as a biosafety level 4 (BSL4) pathogen in most countries and designated a priority pathogen by the World Health Organization (WHO). To facilitate the preclinical assessment of medical countermeasures, we have established two murine models using C57BL/6J IFNAR-/- mice, which lack the IFN/{beta} receptor, infected with the phylogenetically distinct CCHFV strains Afghanistan09-2990 (Afg09) and Kosovo Hoti (Hoti). Infection with both CCHFV strains in IFNAR-/- mice resulted in significant weight loss, with Afg09 infection leading to more severe clinical disease. Quantitative analysis of viral RNA revealed widespread viral dissemination across multiple organs in both models. Detection of infectious virus varied by organ and strain. These results confirm and extend previous findings, providing a deeper understanding of CCHFV strain-specific pathogenesis in IFNAR-/- mice. Thereby, these mouse models represent valuable tools for the evaluation of antiviral therapeutics and vaccine candidates, enabling the investigation of cross-lineage protection against genetically diverse CCHFV isolates.

microbiology↗

bioRxiv2025

Proteolytic processing of the Marburg virus glycoprotein depends on Sec616β and is required for cell entry

Ebola and Marburg virus (EBOV, MARV) cause severe disease and therapeutic options are urgently needed. The Sec61 translocon facilitates ER import of viral glycoproteins (GPs) and may represent a therapeutic target. Here, we report that the Sec61 subunit Sec61{beta}, although dispensable for GP expression, is required for proteolytic cleavage of MARV- but not EBOV-GP and that an intact furin motif is essential for robust cell entry of Marburg- but not Ebolaviruses. Further, MARV- but not EBOV-GP was cleaved by the furin-related enzyme SKI-1, for which a cleavage motif was identified in silico, and cleavage by SKI-1 was impaired in SEC61B-KO cells. In addition, Sec61{beta} was required for normal N-glycosylation of MARV-GP and mutation of a sequon (N563D) abrogated cleavage. Finally, the absence of Sec61{beta} modestly, and blockade of Sec61 via apratoxin S4 markedly, inhibited EBOV and MARV infection. These results reveal a differential protease dependence of MARV and EBOV and identify Sec61 as a potential therapeutic target. Author summaryThe filoviruses Ebola virus (EBOV) and Marburg virus (MARV) spread from animals to humans and can cause deadly outbreaks. These viruses rely on a surface glycoprotein (GP) for infection, which is processed by the enzyme furin in infected human cells. Cleavage of EBOV-GP was thought to be non-essential for infection. However, using lab models for filovirus entry into cells, we discovered that MARV, unlike EBOV, needs this cleavage step to infect cells efficiently. We also found that the host cell protein Sec61{beta} is necessary for proteolytic processing and glycosylation of MARV-GP but not EBOV-GP. In addition, we showed that another cellular enzyme, SKI-1, can process MARV- but not EBOV-GP. Finally, we found that removing Sec61{beta} or blocking Sec61 activity reduced infection by both viruses. These findings show key differences in how the two viruses interact with host cells and suggest that targeting Sec61 could be a promising new strategy to fight Ebola and Marburg virus infections.

microbiology↗