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- Title
The phenuivirus Toscana virus makes an atypical use of vacuolar acidity to enter host cells.
- Authors
Koch, Jana; Xin, Qilin; Obr, Martin; Schäfer, Alicia; Rolfs, Nina; Anagho, Holda A.; Kudulyte, Aiste; Woltereck, Lea; Kummer, Susann; Campos, Joaquin; Uckeley, Zina M.; Bell-Sakyi, Lesley; Kräusslich, Hans-Georg; Schur, Florian KM.; Acuna, Claudio; Lozach, Pierre-Yves
- Abstract
Toscana virus is a major cause of arboviral disease in humans in the Mediterranean basin during summer. However, early virus-host cell interactions and entry mechanisms remain poorly characterized. Investigating iPSC-derived human neurons and cell lines, we found that virus binding to the cell surface was specific, and 50% of bound virions were endocytosed within 10 min. Virions entered Rab5a+ early endosomes and, subsequently, Rab7a+ and LAMP-1+ late endosomal compartments. Penetration required intact late endosomes and occurred within 30 min following internalization. Virus entry relied on vacuolar acidification, with an optimal pH for viral membrane fusion at pH 5.5. The pH threshold increased to 5.8 with longer pre-exposure of virions to the slightly acidic pH in early endosomes. Strikingly, the particles remained infectious after entering late endosomes with a pH below the fusion threshold. Overall, our study establishes Toscana virus as a late-penetrating virus and reveals an atypical use of vacuolar acidity by this virus to enter host cells. Author summary: Toscana virus (TOSV) is a reemerging sandfly-borne enveloped virus causing neuro-invasive infections in humans. The virus is endemic in the Mediterranean basin, with a potential risk of introduction in northern Europe and Asia. Despite its significance, diagnostics, therapeutics, and research on TOSV have been neglected. Here, we developed accurate, sensitive methods to examine the early stages of TOSV infection in both fixed and live human neurons and cell lines, covering virus binding to fusion. Our results highlight the crucial role of late endosomal maturation in TOSV entry, shedding light on atypical viral fusion mechanisms. Notably, the fusion process of TOSV appears, at least in part, reversible, and the virus depends on progressive acidification in endosomes for activation and penetration, rather than a specific pH threshold. The information gained here lays the basis for future research into entry inhibitors against not only TOSV, but all viruses using similar penetration strategies. Our study also emphasizes that only a synergistic combination of innovative structure-function and cell biology analyses will provide a better understanding of virus fusion in the cellular context.
- Subjects
TUSCANY (Italy); ARBOVIRUS diseases; VIRAL envelopes; MEMBRANE fusion; ACIDITY; ENDOSOMES; PLANT viruses
- Publication
PLoS Pathogens, 2023, Vol 19, Issue 8, p1
- ISSN
1553-7366
- Publication type
Article
- DOI
10.1371/journal.ppat.1011562