Multiple Protein–Protein Interactions Drive the Assembly and Budding of the Chikungunya Virion

The assembly of enveloped viruses is a highly orchestrated process that depends on the coupling of multiple protein–protein interactions within a membrane environment. To gain mechanistic insight into this process, we use Chikungunya virus as a model system to study Alphavirus assembly, focusing on the interplay between core–spike and spike–spike interactions. We begin with coarse-grained molecular dynamics simulations to systematically explore how the symmetry of the nucleocapsid core, together with the relative strengths of spike–core and spike–spike interactions, influences budding efficiency and the emergence of icosahedral particle symmetry. Building on these computational results, we performed site-directed mutagenesis on Chikungunya virus 181/25 and examined the consequences for particle assembly and budding in cultured cells, as well as the impact of these mutations during in-cellulo assembly. Our results revealed that canonical core–spike interactions, while necessary, were not sufficient for successful assembly. Instead, lateral interactions among glycoproteins emerged as critical determinants of efficient budding, particle stability, and the maintenance of icosahedral symmetry. Together, these findings provided an integrated computational and experimental framework for understanding the molecular principles governing Alphavirus assembly.