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Liquid–liquid phase separation underpins the formation of replication factories in rotaviruses

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Geiger, Florian 
Acker, Julia 
Papa, Guido 
Wang, Xinyu 
Arter, William E 


RNA viruses induce the formation of subcellular organelles that provide microenvironments conducive to their replication. Here we show that replication factories of rotaviruses represent proteinRNA condensates that are formed via liquid–liquid phase separation of the viroplasm-forming proteins NSP5 and rotavirus RNA chaperone NSP2. Upon mixing, these proteins readily form condensates at physiologically relevant low micromolar concentrations achieved in the cytoplasm of virus-infected cells. Early infection stage condensates could be reversibly dissolved by 1,6-hexanediol, as well as propylene glycol that released rotavirus transcripts from these condensates. During the early stages of infection, propylene glycol treatments reduced viral replication and phosphorylation of the condensate-forming protein NSP5. During late infection, these condensates exhibited altered material properties and became resistant to propylene glycol, coinciding with hyperphosphorylation of NSP5. Some aspects of the assembly of cytoplasmic rotavirus replication factories mirror the formation of other ribonucleoprotein granules. Such viral RNA-rich condensates that support replication of multi-segmented genomes represent an attractive target for developing novel therapeutic approaches.



RNP granules, biomolecular condensates, microfluidics, viral genome assembly, Animals, Cattle, Cell Line, Cytoplasmic Ribonucleoprotein Granules, Gene Expression Regulation, Viral, Genes, Reporter, Glycols, Green Fluorescent Proteins, HEK293 Cells, Haplorhini, Host-Pathogen Interactions, Humans, Osmolar Concentration, Phosphorylation, Propylene Glycol, Protein Processing, Post-Translational, RNA-Binding Proteins, Rotavirus, Signal Transduction, Viral Nonstructural Proteins, Virus Assembly, Virus Replication

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The EMBO Journal

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EMBO Press
European Research Council (337969)
European Commission Horizon 2020 (H2020) Future and Emerging Technologies (FET) (766972)
European Commission Horizon 2020 (H2020) Marie Sk?odowska-Curie actions (841466)