Rab11B is required for binding and entry of recent H3N2, but not H1N1, influenza A isolates

Turner, A. H.; Jaffrani, S. A.; Kubinski, H. C.; Ajayi, D. P.; Owens, M. B.; McTigue, M. P.; Fanuele, C. D.; Appenzeller, C. L.; Despres, H. W.; Schmidt, M. M.; Crothers, J. W.; Bruce, E. A.

doi:10.1101/2025.04.23.650275

bioRxiv · 10.1101/2025.04.23.650275

Rab11B is required for binding and entry of recent H3N2, but not H1N1, influenza A isolates

Turner, A. H.·Jaffrani, S. A.·Kubinski, H. C.·Ajayi, D. P.·Owens, M. B.·McTigue, M. P.·Fanuele, C. D.·Appenzeller, C. L.·Despres, H. W.·Schmidt, M. M.·Crothers, J. W.·Bruce, E. A.

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Abstract

As an obligate intracellular parasite, influenza A virus (IAV) depends on host proteins to complete several important functions, including trafficking viral proteins throughout the cell. Previous work has established a critical role for the cellular vesicular trafficking protein, Rab11A, in transporting the viral genome segments to the site of budding at the plasma membrane. While the role of Rab11A in IAV assembly is relatively well understood, very little is known about the function of a closely related isoform (Rab11B) during influenza virus infection. We have shown that both Rab11A and Rab11B are required for successful IAV infection by current H1N1 or H3N2 isolates. Cells in which either Rab11A or Rab11B were depleted failed to efficiently produce virus, with significant reductions in infectious titer. Surprisingly, our data revealed that recent (2022) H3N2, but not H1N1, isolates failed to efficiently produce viral proteins in single-cycle infections when Rab11B (but not Rab11A) was depleted. Flow cytometry analysis suggests that the defect in protein production is driven by a reduction in the total number of infected cells, rather than a decrease in viral protein production at the single cell level. Using reverse genetics and 7+1 reassortant viruses we mapped this Rab11B-dependent early defect in recent H3N2 isolates to the HA gene. RT-qPCR analysis of H3N2 virions bound to the cell surface showed a [~]50% decrease in virus binding the surface of cells depleted of Rab11B, but not Rab11A. This data suggests a novel role for Rab11B during viral entry, likely at the stage of viral binding, that is specific to H3N2 isolates.

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Turner, A. H., Jaffrani, S. A., Kubinski, H. C., Ajayi, D. P., Owens, M. B., McTigue, M. P., Fanuele, C. D., Appenzeller, C. L., Despres, H. W., Schmidt, M. M., Crothers, J. W., Bruce, E. A.. 2025-04-24. Rab11B is required for binding and entry of recent H3N2, but not H1N1, influenza A isolates. https://doi.org/10.1101/2025.04.23.650275

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