Norovirus VPg Binds RNA through a Conserved N-Terminal K/R Basic Patch

Area of Science:

• Virology
• Molecular Biology
• Structural Biology

Background:

• The viral protein genome-linked (VPg) is essential for norovirus replication.
• Norovirus VPg proteins possess RNA-binding capabilities, particularly within their N-terminal regions.

Purpose of the Study:

• To investigate the RNA-binding properties of murine norovirus (MNV) VPg.
• To identify the specific amino acid residues responsible for RNA interaction.
• To determine the role of RNA binding in MNV VPg-induced cell cycle arrest.

Main Methods:

• Expression of MNV and human norovirus (HuNV) VPg proteins fused to a spidroin tag in insect cells.
• Electrophoretic mobility shift assays (EMSA) using pentaprobe RNA sequences to assess RNA binding.
• Site-directed mutagenesis of basic residues (lysine/arginine) in the MNV VPg N-terminus.
• Analysis of MNV VPg's effect on cell cycle progression (G0/G1 arrest).

Main Results:

• MNV VPg and HuNV VPg proteins exhibit non-specific RNA binding.
• A basic patch comprising lysine/arginine residues within the first 12 amino acids of MNV VPg is critical for RNA binding.
• Mutation of this basic patch abolishes RNA binding.
• The RNA-binding deficient MNV VPg mutant fails to induce G0/G1 cell cycle arrest.

Conclusions:

• Norovirus VPg proteins utilize a disordered N-terminal basic patch for non-specific RNA binding.
• This RNA interaction is a key mechanism by which MNV VPg influences host cell cycle progression.
• Understanding VPg-RNA interactions offers potential targets for antiviral strategies against noroviruses.