Ling-Dong Xu, Fei Zhang, Chao Miao, Xinyuan Yu, Yezhang Zhu, Mengdi Zhang, Shengduo Liu, Siddharth Sridhar, Qiming Sun, Dante Neculai, Qi Zhang, Li Shen, Tingbo Liang, Cunqi Ye, Yao‐Wei Huang, Pinglong Xu
Lipid droplets (LDs) are dynamic organelles that mediate lipid metabolism and various cellular processes. However, the interplay between hepatocyte LDs and hepatitis E remains poorly understood. Using targeted lipidomics and lipid profiling in rodent models, we reveal that hepatitis E virus (HEV) infection substantially increases hepatic LD biogenesis. Mechanistically, HEV pORF3 is a key LD biogenesis inducer and an essential factor for viral infectivity in vivo. pORF3 undergoes liquid-liquid phase-separation to form condensates that associate with LD phospholipid monolayer peripherally to upregulate cholesterol anabolic pathways, thereby promoting triacylglycerol and cholesterol ester synthesis. Consistently, genetic loss of ORF3 or pharmacologic reduction of LD biogenesis with the statin atorvastatin substantially suppressed HEV infection in vivo. These findings identify LD biogenesis as a host dependency for HEV infectivity and propose alternative strategies for HEV intervention by targeting LD-directed metabolic pathways.