Regulation of Influenza A virus intracellular life cycle by cellular E3-ubiquitin ligase via the ubiquitination of viral and cellular proteins
Jingshu Zhang1,2, Elise Biquand2, Mart Lamers3, Marwah Karim2, Ying Fan1, Caroline Demeret2, Sumana Sanyal1,4 1HKU-Pasteur Research Pole, School of Public Health and 4School of Biomedical Sciences, The University of Hong Kong, Hong Kong, China; 2Molecular Genetics of RNA Viruses Unit, Department of Virology, Institut Pasteur, Paris, France; 3Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands
Post-translational modifications of viral proteins and cellular factors are key strategies to regulate viral infection. We have previously reported that one such factor, TSG101, is required for intracellular trafficking of the Influenza A virus (IAV) Hemagglutinin and is effectively regulated by ISGylation. In this context, we employed proteomic strategies to identify interactors of TSG101 that operate during IAV infection. In parallel, we screened for host ubiquitin proteasome system (UPS) components, which target IAV polymerase subunit PB2 to modulate viral infection. We characterized an E3-ubiquitin ligase, MGRN1, that was identified from both screens and appeared to critically regulate IAV replication and production. MGRN1, the known E3-ligase for TSG101, was highly enriched in our interactome data from IAV-infected samples compared to mock-treated, but not in the interferon-treated samples. MGRN1 bound to PB2 from different strains of IAV regardless of their pathogenicity. In addition, MGRN1 enhanced ubiquitination of PB2 - an effect that was attenuated by the presence of TSG101. Loss of MGRN1 resulted in attenuated virion release. The subcellular distribution of MGRN1 shifted from the nucleus to the cytosol upon IAV infection, suggesting that MGRN1 was hijacked by IAV to facilitate both viral replication and assembly. We hypothesize that MGRN1 uses both IAV PB2 and TSG101, which probably compete as its substrates for ubiquitination. MGRN1 therefore appears to function as the intermediary between replication and assembly of IAV, mediating different stages of the viral intracellular life cycle. Our data implicates a host factor in the IAV life cycle that has the potential for being targeted by chemical inhibitors to attenuate IAV infection. This study will also contribute to a fundamental understanding of the interplay between UPS and the innate immune system during viral infection.
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