The Src family kinase c-Yes is required for maturation of West Nile virus particles

Area of Science:

• Virology
• Cellular Biology
• Molecular Biology

Background:

• The cellular mechanisms governing West Nile virus (WNV) replication are not fully elucidated.
• Src family kinases (SFKs) are implicated in various cellular processes, but their role in WNV infection remains unclear.

Purpose of the Study:

• To investigate the role of cellular genes, specifically SFKs, in WNV replication.
• To determine the specific SFK involved in WNV production and identify its mechanism of action.

Main Methods:

• Analysis of cellular transcripts during WNV infection.
• Treatment of WNV-infected cells with SFK inhibitor PP2.
• RNA interference (RNAi) to knock down c-Yes and c-Src expression.
• Western blotting and endoglycosidase H (endoH) digestion to assess viral protein processing.
• Electron microscopy to visualize virion localization.

Main Results:

• WNV infection upregulated the expression of c-Yes, an SFK.
• Inhibition of SFK activity with PP2 or c-Yes knockdown significantly reduced WNV titers.
• PP2 treatment did not affect intracellular viral RNA or protein levels, indicating no impact on early replication.
• PP2 treatment inhibited the acquisition of endoH-resistant glycans by the WNV E glycoprotein, suggesting impaired trafficking beyond the ER.
• Electron microscopy revealed accumulation of virions within the ER of PP2-treated cells.

Conclusions:

• SFK activity, particularly that of c-Yes, is essential for WNV replication.
• c-Yes plays a critical role in the post-assembly stages of WNV infection, specifically in virion egress.
• Inhibition of SFK activity prevents WNV virion transit through the secretory pathway, trapping them in the endoplasmic reticulum.