Gonzalez-Orozco, M. | bioRxiv Science

bioRxiv2024

TRIM7 ubiquitinates SARS-CoV-2 membrane protein to limit apoptosis and viral replication

SARS-CoV-2 is a highly transmissible virus that causes COVID-19 disease. Mechanisms of viral pathogenesis include excessive inflammation and viral-induced cell death, resulting in tissue damage. We identified the host E3-ubiquitin ligase TRIM7 as an inhibitor of apoptosis and SARS-CoV-2 replication via ubiquitination of the viral membrane (M) protein. Trim7-/- mice exhibited increased pathology and virus titers associated with epithelial apoptosis and dysregulated immune responses. Mechanistically, TRIM7 ubiquitinates M on K14, which protects cells from cell death. Longitudinal SARS-CoV-2 sequence analysis from infected patients revealed that mutations on M-K14 appeared in circulating variants during the pandemic. The relevance of these mutations was tested in a mouse model. A recombinant M- K14/K15R virus showed reduced viral replication, consistent with the role of K15 in virus assembly, and increased levels of apoptosis associated with the loss of ubiquitination on K14. TRIM7 antiviral activity requires caspase-6 inhibition, linking apoptosis with viral replication and pathology.

microbiology↗

bioRxiv2023

Ebola Virus VP35 Interacts Non-Covalently with Ubiquitin Chains to Promote Viral Replication Creating New Therapeutic Opportunities

Ebolavirus (EBOV) belongs to a family of highly pathogenic viruses that cause severe hemorrhagic fever in humans. EBOV replication requires the activity of the viral polymerase complex, which includes the co-factor and Interferon antagonist VP35. We previously showed that the covalent ubiquitination of VP35 promotes virus replication by regulating interactions with the polymerase complex. In addition, VP35 can also interact non-covalently with ubiquitin (Ub); however, the function of this interaction is unknown. Here, we report that VP35 interacts with free (unanchored) K63-linked polyUb chains. Ectopic expression of Isopeptidase T (USP5), which is known to degrade unanchored polyUb chains, reduced VP35 association with Ub and correlated with diminished polymerase activity in a minigenome assay. Using computational methods, we modeled the VP35-Ub non-covalent interacting complex, identified the VP35-Ub interacting surface and tested mutations to validate the interface. Docking simulations identified chemical compounds that can block VP35-Ub interactions leading to reduced viral polymerase activity that correlated with reduced replication of infectious EBOV. In conclusion, we identified a novel role of unanchored polyUb in regulating Ebola virus polymerase function and discovered compounds that have promising anti-Ebola virus activity. Significance StatementEbola virus infection can result in high mortality rates with extreme risk of person-to-person transmission. Sporadic outbreaks in Africa have resulted in thousands of fatal cases, highlighting that there is still insufficient knowledge to develop effective antiviral therapies. Like other viruses, Ebola utilizes the host machinery to replicate. Understanding how viral and host proteins interact can help identifying targets for the rational design of antiviral drugs. Here, we identified interactions between the cellular ubiquitin machinery and the Ebola virus polymerase cofactor protein VP35, which are important for efficient virus replication. We developed an approach to identify and block these virus-host interactions using small chemical compounds, which provides a useful tool to study functional molecular mechanisms and at the same time a potential approach to antiviral therapies.

immunology↗