Selenoprotein S Interacts with the Replication and Transcription Complex of SARS-CoV-2 by Binding nsp7.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) replicates and evades detection using ER membranes and their associated protein machinery. Among these hijacked human proteins is selenoprotein S (selenos). This selenoprotein takes part in the protein quality control, signaling, and the regulation of cytokine secretion. While the role of selenos in the viral life cycle is not yet known, it has been reported to interact with SARS-CoV-2 nonstructural protein 7 (nsp7), a viral protein essential for the replication of the virus. We set to study whether selenos and nsp7 interact directly and if they can still bind when nsp7 is bound to the replication and transcription complex of the virus. Using biochemical assays, we show that selenos binds directly to nsp7. In addition, we found that selenos can bind to nsp7 when it is in a complex with the coronavirus's minimal replication and transcription complex, comprised of nsp7, nsp8, and the RNA-dependent RNA polymerase nsp12. In addition, through crosslinking experiments, we mapped the interaction sites of selenos and nsp7 in the replication complex and showed that the hydrophobic segment of selenos is essential for binding to nsp7. This arrangement leaves an extended helix and the intrinsically disordered segment of selenos-including the reactive selenocysteine-exposed and free to potentially recruit additional proteins to the replication and transcription complex.

Selenoprotein S Binds to the SARS‐CoV‐2 Replication Complex

The severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2) uses endoplasmic reticulum membranes and associated human proteins for its replication and to evade detection. One human protein recruited by several viral proteins is selenoprotein S, which takes part in the endoplasmic reticulum protein degradation pathway, NFkB signaling, and cytokines secretion. A key interaction was reported with SARS‐CoV‐2 non‐structural protein (nsp7), which is essential for virus replication. However, it was unclear whether selenoprotein S and nsp7 interact directly and whether the interaction is possible when nsp7 forms a complex with the other components of the virus's replication machinery. We used biochemical assays to show that selenoprotein S binds nsp7, including when nsp7 is in complex with the coronavirus's RNA‐dependent RNA polymerase. This places selenoprotein S at the heart of the coronavirus's replication complex and marks it as the first human protein shown to directly interact with the viral replication complex. Cross‐linking experiments were employed to map the interactions of selenoprotein S and nsp7 in the replication complex. We show that the hydrophobic segment of selenoprotein S is essential for binding nsp7. This arrangement leaves an extended helix and the intrinsically disordered region of selenoprotein S exposed and free to recruit additional proteins to the complex.

The role of human selenoprotein S in SARS-CoV-2 replication

COVID-19, caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), spread quickly all over the world and caused a pandemic that urged the scientific community to study the virus. Selenoprotein S is an endoplasmic reticulum (ER) membrane associated selenoprotein that is involved in protein degradation and NF?? signaling. It also contributes to reducing proinflammatory cytokines TNF-α, IL-1? and IL-6. Selenoprotein S is known to interact with SARS-CoV-2 non-structural protein (nsp7), which is part of the virus replication machine. However, it is not clear whether they interact directly and whether selenoprotein S can still bind nsp7 once it is in complex with the virus? replication machinery. The role of selenoprotein S in the replication complex is unknown. We used biochemical assays to investigate these open questions and to identify which segment of selenoprotein S is involved in the interaction and which segment is free to recruit other human or viral proteins. We show that selenoprotein S binds tightly and directly to nsp7. Selenoprotein S is a membrane protein and its hydrophobic region is the main segment involved in binding nsp7. The soluble cytosolic segment cannot bind to nsp7. Therefore, it is free for binding to other protein partners. In addition, selenoprotein S was able to bind the nsp7 and nsp8 complex suggesting that it can bind the core replication complex directly. Thus, we suggest that selenoprotein S is a component of the virus core replication complex and may be responsible for recruiting other human proteins to the replication complex.