PACT inhibits the replication of SARS-CoV-2 through the blockage of GSK-3β-N-nsp3 cascade.

The protein activator of protein kinase R (PKR) (PACT) has been shown to play a crucial role in stimulating the host antiviral response through the activation of PKR, retinoic acid-inducible gene I, and melanoma differentiation-associated protein 5. Whether PACT can inhibit viral replication independent of known mechanisms is still unrevealed. In this study, we show that, like many viruses, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) hijacks GSK-3β to facilitate its replication. GSK-3β-induced phosphorylation on N protein increased the interaction between N protein and nsp3. Thus, GSK-3β-N-nsp3 cascade promotes viral replication. Although SARS-CoV-2 can sabotage the activation of AKT, the upstream proteins suppressing the activation of GSK-3β, we found that the host can use PACT, another protein kinase, instead of AKT to decrease the activity of GSK-3β and the interaction between PACT and GSK-3β is enhanced upon viral infection. Moreover, PACT inhibited the activity of GSK-3β independent of its well-studied double-stranded RNA binding and PKR activating ability. In summary, this study identified an unknown function of PACT in inhibiting SARS-CoV-2 replication through the blockage of GSK-3β-N-nsp3 cascade.