Evolution of a fuzzy ribonucleoprotein complex in viral assembly

Area of Science:

• Virology
• Biophysics
• Molecular Biology

Background:

• The SARS-CoV-2 nucleocapsid (N) protein plays a crucial role in viral assembly by condensing viral RNA into ribonucleoprotein particles (RNPs).
• Previous work demonstrated that N protein mutations can alter its biophysical properties and self-association.
• High-resolution structures of RNPs are lacking, necessitating biochemical and biophysical approaches to understand their architecture.

Purpose of the Study:

• To investigate the impact of specific N-protein mutations found in SARS-CoV-2 variants of concern on RNP formation.
• To elucidate how these mutations affect the stability and assembly dynamics of RNPs.
• To understand the role of RNP complex variability in viral fitness and host adaptation.

Main Methods:

• Biophysical techniques were employed to study RNP formation.
• A virus-like particle assay was utilized to assess viral assembly.
• Reverse genetics experiments were conducted to evaluate the functional consequences of N-protein mutations.

Main Results:

• Convergent evolution was observed, with independent mutations strengthening existing binding interfaces to compensate for those that decrease RNP stability.
• The P13L mutation, characteristic of Omicron variants, was found to enhance RNP assembly and increase viral fitness.
• SARS-CoV-2 RNP complexes exhibit significant variability in sequence, conformation, and thermodynamic/kinetic stability.

Conclusions:

• N-protein mutations in SARS-CoV-2 variants can significantly alter RNP assembly and stability, impacting viral fitness.
• The Omicron variant's P13L mutation enhances RNP formation, contributing to increased viral replication and host adaptation.
• The pleomorphic nature of RNPs, characterized by fuzzy clusters and weak binding interfaces, likely optimizes reversible RNA condensation for viral survival and evolution.