Single-Molecule Dynamics of SARS-CoV-2 5' Cap Recognition by Human eIF4F

Area of Science:

• Molecular Biology
• Virology
• Biochemistry

Background:

• Coronaviruses, including SARS-CoV-2, rely on the eIF4F translation factor complex to initiate viral RNA translation via cap recognition.
• The precise coordination of eIF4F subunits (eIF4E, eIF4G, eIF4A) during translation initiation and their impact on cap-binding efficiency remain incompletely understood.
• Modulating eIF4F function is a potential strategy for inhibiting coronavirus replication.

Approach:

• Utilized a single-molecule fluorescence assay to monitor the interaction between the eIF4E cap-binding protein and the SARS-CoV-2 5'-UTR.
• Investigated the roles of individual eIF4F subunits (eIF4A, eIF4G) and the complete eIF4F complex in cap recognition.
• Assessed the effect of rocaglamide, an eIF4A inhibitor, on viral translation and the influence of eIF4E phosphorylation on cap-binding.

Key Points:

• Free eIF4A enhances the accessibility of the RNA cap for eIF4E binding, while eIF4G alone does not significantly alter eIF4E-RNA interaction kinetics.
• The formation of the complete eIF4F complex markedly changes eIF4E-cap interactions, indicating that coordinated eIF4E and eIF4A activities determine overall cap recognition efficiency.
• The phosphomimetic mutant eIF4E(S209D) exhibits enhanced binding to the viral 5'-UTR compared to wild-type eIF4E, suggesting a role for phosphorylation in regulating cap recognition.

Conclusions:

• The study elucidates the dynamic interplay between eIF4F subunits and viral RNA during translation initiation.
• Coordinated action of eIF4E and eIF4A is crucial for efficient cap recognition on the SARS-CoV-2 5'-UTR.
• Findings provide insights into the mechanisms of viral translation and potential targets for antiviral drug development.