I'm very excited to share the latest work from my group, led by @ahturner934.bsky.social. We revisited one of my favorite proteins (Rab11) and its role in influenza infection& discovered surprising new things. Rab11B is required for entry of current H3N2 isolates! 1/n www.biorxiv.org/content/10.1... - ThreadSky

brucelab.bsky.social • 21 days ago

I'm very excited to share the latest work from my group, led by @ahturner934.bsky.social. We revisited one of my favorite proteins (Rab11) and its role in influenza infection& discovered surprising new things. Rab11B is required for entry of current H3N2 isolates! 1/n https://www.biorxiv.org/content/10.1101/2025.04.23.650275v1

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brucelab.bsky.social•21 days ago

We started by depleting two Rab11 isoforms (A and B), separately or together (panel A), in human lung cells then infecting with recent H1N1 (panel B) or H3N2 (panel C) subtypes we isolated from VT clinical samples. Both Rab11 isoforms were required for efficient production of infectious virus. 2/n

brucelab.bsky.social•21 days ago

Rab11 is well known to transport the genome segments late in IAV infection, but has not been reported to play a role prior to protein production. Which is why we were very surprised to see that cells lacking Rab11B and infected with the H3N2 isolate failed to efficiently make viral proteins. 3/n

brucelab.bsky.social•21 days ago

To make sure this wasn't an artifact of our particular H3N2, we repeated this experiment with a second H3N2 isolate from a different state (thanks to @andrewpekosz.bsky.social)! The 2nd virus showed the same phenotype- both Rab11s required for virus production, Rab11B needed early in infection. 4/n

brucelab.bsky.social•21 days ago

Cells infected with the H1N1 isolate made viral proteins just fine, suggesting the defect in infectious virion production happened late in infection (fitting with Rab11A's well characterized role in transporting the RNPs to the site of budding). 5/n

brucelab.bsky.social•21 days ago

Since western blots measure bulk protein expression, and transient knockdowns may not reach 100% of cells, we wanted to see what was happening at a single cell level. Flow cytometry showed that most Rab11B- cells failed to produce any viral proteins, rather than a global decrease in translation. 6/n

brucelab.bsky.social•21 days ago

We used an NH4Cl time of addition experiment to prevent endosomal acidification (and thus viral fusion) to map the kinetics of viral entry. Rab11B- cells had significantly decreased rates of entry, but the kinetics of internalization was not delayed (suggesting the problem was further upstream). 7/n

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