Abstract

Influenza A virus has an eight-partite RNA genome that during viral assembly forms a supramolecular complex containing one copy of each RNA. Genome assembly is a selective process driven by RNA-RNA interactions and is thought to lead to discrete punctate structures scattered through the cytosol. Here, we show that contrary to the accepted view, formation of these structures is not dependent on RNA-RNA interactions among distinct viral ribonucleoproteins (vRNPs), as they assemble in cells expressing only one vRNP type. We demonstrate that these viral inclusions display characteristics of liquid organelles, segregating from the cytosol without a delimitating membrane, dynamically exchanging material, deforming easily and adapting fast to hypotonic shock. We provide evidence that they develop close to the Endoplasmic Reticulum Exit Sites (ERES), being dependent on continuous ER-Golgi vesicular cycling. We show that viral inclusions do not promote escape to interferon response, and propose that they facilitate selected RNA-RNA interactions in a liquid environment of concentrated vRNPs.