Abstract
ABSTRACT
Bat-origin swine acute diarrhea syndrome coronavirus (SADS-CoV) causes diarrhea and death in piglets and also poses a cross-species transmission risk due to its broad host cell tropism. In the absence of commercial vaccines or therapeutics, identifying the host factors exploited by SADS-CoV may provide potential targets for antiviral therapies. Inhibition of glycogen synthase kinase 3 (GSK3) restricts diverse coronavirus replication by targeting the N protein and disrupting its phosphorylation. Here, we identified GSK3 as a crucial host factor for SADS-CoV RNA synthesis. Both GSK3 deficiency and treatment with a classical GSK3 inhibitor, lithium chloride (LiCl), strongly impaired SADS-CoV dsRNA synthesis. LiCl treatment did not alter electrophoretic mobility or expression of SADS-CoV N protein in N-transfected Huh7 cells but significantly reduced NSP12 protein levels. The absence of GSK3 led to the proteasomal degradation of NSP12 to reduce viral RNA synthesis. However, GSK3 interacted with ubiquitin-specific peptidase 27X (USP27X) and promoted the USP27X-mediated deubiquitination of NSP12, thereby enhancing NSP12 stabilization and facilitating viral RNA synthesis. LiCl not only protected neonatal mice from lethal SADS-CoV challenge but also exhibited a notable broad-spectrum antiviral effect against PEDV and TGEV. Our findings demonstrate that GSK3-mediated NSP12 stabilization is essential for SADS-CoV RNA synthesis, underscoring the potential of GSK3 as a broad-spectrum antiviral target for emerging and re-emerging swine coronaviruses.
IMPORTANCE
Zoonotic transmission of coronaviruses continues to pose a significant threat to human populations. Swine acute diarrhea syndrome coronavirus (SADS-CoV) has been identified as a potentially high-risk emerging coronavirus pathogen that could adversely affect the global economy and human health. However, the replication mechanisms of SADS-CoV remain largely unexplored. Our study demonstrates that GSK3 stabilizes NSP12 by interacting with USP27X, thereby enhancing the synthesis of SADS-CoV dsRNA. LiCl, a GSK3 inhibitor, exhibits broad-spectrum antiviral activity against swine enteric alphacoronaviruses. Our work is distinguished for the first time by the GSK3–USP27X regulatory axis that controls the stability of NSP12 and indicates that GSK3 is a promising therapeutic target for the development of broad-spectrum antiviral drugs against emerging or re-emerging swine coronaviruses.