ARF-like GTPase 8B orchestrates lipophagy and exocytosis to drive single-stranded RNA virus replication

Single-stranded RNA (ssRNA) viruses, including foot‑and‑mouth disease virus (FMDV), enterovirus 71 (EV71), and vesicular stomatitis virus (VSV), reprogram host lipid metabolism to facilitate replication. However, the mechanisms governing virus‑induced lipid droplet (LD) degradation remain poorly defined, which limits the development of host‑directed antiviral strategies.

We employed virological, biochemical, and imaging approaches to investigate the role of the lysosomal GTPase ARL8B in ssRNA virus infection. Lipid droplet dynamics, lipophagy, and lysosomal exocytosis were assessed using confocal microscopy and biochemical assays. Small‑molecule inhibitors targeting ARL8B were identified through virtual screening and validated in vitro for antiviral activity. The in vivo efficacy of selected inhibitors was evaluated in a murine EV71 infection model, with disease severity and survival as endpoints.

ARL8B was identified as a central host factor that promotes replication of multiple ssRNA viruses by coordinating LD degradation with viral egress. Mechanistically, ARL8B drove lysosome‑dependent LD breakdown via selective lipophagy, releasing free fatty acids that fuel membrane remodeling within viral replication complexes. Concurrently, ARL8B facilitated viral exit through a noncanonical lysosomal-exosome pathway, enhancing progeny virus dissemination. Virtual screening yielded small‑molecule ARL8B inhibitors that potently suppressed ssRNA virus replication in vitro. In a murine EV71 infection model, these inhibitors provided robust protection and substantially reduced disease severity.

Our findings establish ARL8B as a key host factor coupling lipid catabolism with viral export, and unveil a class of broad‑spectrum antiviral candidates targeting a lipid reprogramming node essential for multiple pathogenic RNA viruses. This study provides a mechanistic framework for developing host‑directed antiviral therapies against ssRNA viruses.