Discerning the mechanisms of cell membrane budding by coronavirus non-structural proteins nsp3, nsp4, and nsp6

The occurrence of viral infections in human populations dates to the early 20th century and a large variety of viral pathogens have emerged since. Specifically, Coronaviridae have caused the severe acute respiratory syndrome (SARS) epidemic, the Middle-East respiratory syndrome (MERS), and the SARS-CoV-2 pandemic. Throughout coronaviral infections in host cells, a set of non-structural proteins (nsps) is crucial for facilitating viral replication - nsp3, nsp4, and nsp6. These membrane-spanning proteins were determined to play an active role in deforming the endoplasmic reticulum (ER) membrane in conjunction with other host proteins to form double-membrane vesicles (DMV). DMVs are important replicative structures, which contribute to viral RNA synthesis by providing an ideal environment of cellular and viral constituents while offering protection from host cell defence mechanisms. Despite the formation of DMV structures induced by nsp proteins being conserved in coronaviruses and the SARS epidemic dating back a decade, essential biochemical and biophysical mechanisms governing nsp-induced DMV formation remain poorly understood. Thus, we propose the use of reconstituted biomimetic membrane systems and SARS-CoV-2-derived nsp proteins and peptides to study the biochemical and biomechanical aspects underlying DMV formation. This can be achieved by determining how biophysical properties and interaction dynamics of lipid membranes are altered when bound to nsps, by means of a wide range of biophysical and imaging methods. The results of this project will greatly further our understanding of the molecular mechanisms in nsp-driven coronaviral replication.