Irreversible furin cleavage site exposure renders immature tick-borne flaviviruses fully infectious

• Published in: Nature communications Vol. 16; no. 1; pp. 7491 - 17
• Main Authors: Holoubek, Jiří, Salát, Jiří, Matkovic, Milos, Bednář, Petr, Novotný, Pavel, Hradilek, Martin, Majerová, Taťána, Rosendal, Ebba, Eyer, Luděk, Fořtová, Andrea, Beránková, Michaela, Bell-Sakyi, Lesley, Överby, Anna K., Cavalli, Andrea, Bonomi, Massimiliano, Rey, Félix A., Růžek, Daniel
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 12.08.2025; Nature Publishing Group; Nature Portfolio
• Subjects: 13/1; 13/106; 13/51; 14/1; 14/63; 631/326/596/2555; 631/326/596/2557; 64; 64/60; Animals; Aquatic insects; Arachnids; Cleavage; Dengue fever; Dimers; Encephalitis; Encephalitis Viruses, Tick-Borne - genetics; Encephalitis Viruses, Tick-Borne - metabolism; Encephalitis Viruses, Tick-Borne - pathogenicity; Encephalitis Viruses, Tick-Borne - physiology; Endoplasmic reticulum; Experiments; Exposure; Female; Flaviviridae; Flavivirus - genetics; Flavivirus - pathogenicity; Flavivirus - physiology; Furin; Furin - genetics; Furin - metabolism; Genomes; Glycoproteins; Golgi apparatus; Humanities and Social Sciences; Humans; Hydrogen-Ion Concentration; Infections; Infectivity; Maturation; Mice; Mice, Inbred BALB C; Modelling; Molecular dynamics; Mosquitoes; multidisciplinary; pH effects; Phenotypes; Proteins; Proteolysis; Science; Science (multidisciplinary); Site-directed mutagenesis; Tick-borne encephalitis; Vector-borne diseases; Viral Envelope Proteins - chemistry; Viral Envelope Proteins - genetics; Viral Envelope Proteins - metabolism; Virions; Virus Assembly; Viruses; West Nile virus
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-025-62750-6

Flavivirus assembly is driven by the envelope glycoproteins pre-membrane (prM) and envelope (E) in the neutral pH environment of the endoplasmic reticulum. Newly budded, spiky particles are exported through the Golgi apparatus, where mildly acidic pH induces a major surface rearrangement. The glycoproteins reorganize into (prM/E)\₂ complexes at the surface of smooth particles, with prM trapped at the E dimer interface, thereby exposing a furin cleavage site (FCS) for proteolytic maturation into infectious virions. Here, we show that in the absence of furin, immature tick-borne flavivirus particles—tick-borne encephalitis virus, Langat virus, and Louping ill virus—remain fully infectious and pathogenic in female BALB/c mice, in contrast to mosquito-borne flaviviruses such as Usutu, West Nile, and Zika viruses. We further show that the FCS in tick-borne viruses remains exposed at neutral pH, allowing furin at the surface of target cells to activate viral fusogenicity, while mosquito-borne counterparts require acidic re-exposure. Mutations increasing the dynamic behavior of the E dimer mimic the mosquito-borne phenotype, with retracted FCS at neutral pH and loss of infectivity. Our multidisciplinary approach—combining virological assays, targeted mutagenesis, structural modeling, and molecular dynamics simulations—highlights the role of E dimer dynamics in regulating flavivirus maturation and infectivity. Immature particles of flaviviruses transmitted by mosquitoes are known to be non-infectious. Here, the authors show that immature tick-borne flaviviruses stay infectious in vivo and using structural modeling they resolve unique structural features that distinguish tick-borne from mosquito-borne flaviviruses.