The role of NSP6 in the biogenesis of the SARS-CoV-2 replication organelle

• Published in: Nature (London) Vol. 606; no. 7915; pp. 761 - 768
• Main Authors: Ricciardi, Simona, Guarino, Andrea Maria, Giaquinto, Laura, Polishchuk, Elena V., Santoro, Michele, Di Tullio, Giuseppe, Wilson, Cathal, Panariello, Francesco, Soares, Vinicius C., Dias, Suelen S. G., Santos, Julia C., Souza, Thiago M. L., Fusco, Giovanna, Viscardi, Maurizio, Brandi, Sergio, Bozza, Patrícia T., Polishchuk, Roman S., Venditti, Rossella, De Matteis, Maria Antonietta
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 23-06-2022; Nature Publishing Group
• Subjects: 13; 13/1; 13/106; 13/109; 13/89; 14; 14/19; 14/28; 14/33; 14/34; 14/35; 631/80/313/1463; 631/80/642/1463; 82; 82/29; 82/80; Amino acids; Antiviral agents; Biological activity; Biosynthesis; Clusters; Connectors; Coronaviridae; Coronaviruses; COVID-19; Endoplasmic reticulum; Humanities and Social Sciences; Lipids; Membrane vesicles; Membranes; Microscopy; multidisciplinary; Mutants; Mutation; Oligomerization; Proteins; Refurbishment; Replication; Science; Science (multidisciplinary); Severe acute respiratory syndrome coronavirus 2; Structural proteins; Tethering; Zippers
• ISSN: 0028-0836; 1476-4687
• DOI: 10.1038/s41586-022-04835-6

SARS-CoV-2, like other coronaviruses, builds a membrane-bound replication organelle to enable RNA replication
1
. The SARS-CoV-2 replication organelle is composed of double-membrane vesicles (DMVs) that are tethered to the endoplasmic reticulum (ER) by thin membrane connectors
2
, but the viral proteins and the host factors involved remain unknown. Here we identify the viral non-structural proteins (NSPs) that generate the SARS-CoV-2 replication organelle. NSP3 and NSP4 generate the DMVs, whereas NSP6, through oligomerization and an amphipathic helix, zippers ER membranes and establishes the connectors. The NSP6(ΔSGF) mutant, which arose independently in the Alpha, Beta, Gamma, Eta, Iota and Lambda variants of SARS-CoV-2, behaves as a gain-of-function mutant with a higher ER-zippering activity. We identified three main roles for NSP6: first, to act as a filter in communication between the replication organelle and the ER, by allowing lipid flow but restricting the access of ER luminal proteins to the DMVs; second, to position and organize DMV clusters; and third, to mediate contact with lipid droplets (LDs) through the LD-tethering complex DFCP1–RAB18. NSP6 thus acts as an organizer of DMV clusters and can provide a selective means of refurbishing them with LD-derived lipids. Notably, both properly formed NSP6 connectors and LDs are required for the replication of SARS-CoV-2. Our findings provide insight into the biological activity of NSP6 of SARS-CoV-2 and of other coronaviruses, and have the potential to fuel the search for broad antiviral agents.
The non-structural protein NSP6 in SARS-CoV-2 has a key role in viral replication by zippering the endoplasmic reticulum membrane to establish connectors between the double-membrane vesicles of the viral replication organelle and the endoplasmic reticulum.