Comprehensive transcriptomic analysis identifies cholesterol transport pathway as a therapeutic target of porcine epidemic diarrhea coronavirus

Porcine epidemic diarrhea virus (PEDV) is a highly contagious virus that poses a serious threat to the global pig industry. Despite extensive efforts, the mechanism underlying virus entry for PEDV remains elusive. In this study, we first identified PEDV-susceptible and non-susceptible cell lines by using PEDV spike pseudotyped vesicular stomatitis virus. Subsequently, we conducted a comprehensive transcriptomic analysis on these cell lines. Through integrating differential expression gene analysis with weighted gene co-expression network analysis, we identified the key pathways that are correlated with the PEDV entry. Our analysis revealed a strong correlation between cholesterol, sterols, and lipid transport with PEDV entry, suggesting a potential role for cholesterol transport in the PEDV entry. For further investigation, we treated Huh7, Vero and LLC-PK1 cells with a cholesterol transport inhibitor, ezetimibe, and observed a significant inhibition of PEDV entry and subsequent viral replication in these cells. Interestingly, pre-treating Huh7 cells with ezetimibe resulted in an increase in the entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Middle East respiratory syndrome coronavirus (MERS-CoV) pseudoviruses. Moreover, we found that cholesterol could facilitate the entry of PEDV into Huh7 and Vero cells, and this promoting effect can be blocked by ezetimibe. These findings suggest that targeting cholesterol transport specifically inhibits PEDV entry into susceptible cells. Our study offers novel insights into the mechanism of PEDV entry and the development of new therapeutic strategies against this economically important virus.