Tombusviruses orchestrate the host endomembrane system to create elaborate membranous replication organelles.

Positive-strand RNA viruses depend on intensive manipulation of subcellular organelles and membranes to create unique viral replication organelles (VROs), which represent the sites of robust virus replication. The host endomembrane-based protein-trafficking and vesicle-trafficking pathways are specifically targeted by many (+)RNA viruses to take advantage of their rich resources. We summarize the critical roles of co-opted endoplasmic reticulum subdomains and associated host proteins and COPII vesicles play in tombusvirus replication. We also present the surprising contribution of the early endosome and the retromer tubular transport carriers to VRO biogenesis. The central player is tomato bushy stunt virus (TBSV), which provides an outstanding system based on the identification of a complex network of interactions with the host cells. We present the emerging theme on how TBSV uses tethering and membrane-shaping proteins and lipid modifying enzymes to build the sophisticated VRO membranes with unique lipid composition.