Papain‐Like Protease of SARS‐CoV‐2 Induces Intestinal Inflammation via the ISG15 Pathway: Identification of Natural Compound Inhibitors

ABSTRACT The SARS‐CoV‐2 papain‐like protease (PLpro) is a multifunctional viral protein that facilitates viral assembly and disrupts host immune responses by removing interferon‐stimulated gene 15 (ISG15) modifications from target proteins. However, the mechanisms by which PLpro‐mediated immune evasion leads to inflammatory responses are not well understood. In this study, we demonstrate for the first time that PLpro induces inflammation in colonic epithelial cells and colonic inflammation in a mouse model, acting through the ISG15 signaling pathway. Using high‐throughput screening, we identified two natural product‐derived compounds, coptisine sulfate and (+)‐shikonin, that inhibit the interaction between PLpro and ISG15 at the PLpro/ISG15 interface. We further investigated the mechanism of action of these two inhibitors using surface plasmon resonance, thermal shift assays, molecular docking, and molecular simulation studies. Importantly, both coptisine sulfate and (+)‐shikonin were able to reduce intestinal inflammation in the PLpro‐induced mouse model. These findings provide novel insights into how the SARS‐CoV‐2 PLpro can drive inflammatory responses and introduce two natural compound‐based inhibitors that may offer a new approach to alleviate gastrointestinal complications associated with COVID‐19 infection. Our results highlight the potential of targeting the PLpro/ISG15 interaction as a therapeutic strategy against SARS‐CoV‐2.