Fig. 4

The interconnection of canonical and extracellular vesicle-mediated HCV life cycle. A The schematic outlines the genome structure of HCV, encompassing ten viral proteins expressed through a polyprotein via an internal ribosome entry site (IRES). B The canonical HCV life cycle initiates with binding and attachment to receptors such as LDLR, CD81, and SCARB1 on cells, facilitating entry into hepatocytes. Following initial attachment, claudin-1 (CLDN1) and Occludin (OCLN) mediate translocation to tight junctions and promote Clathrin/dynamin-mediated endocytosis, with the involvement of Rab5. After Rab7-mediated uncoating, viral genomic RNA is released for translation and RNA replication, facilitated by Rab9. Polyprotein translation begins in the endoplasmic reticulum (ER), where ribosomal subunits bind to HCV-RNA. After proteolysis, mature viral proteins induce the rearrangement of host cell membranes, forming double-membrane vesicles within the membranous web. The NS5B RNA-dependent RNA polymerase catalyzes negative-sense RNA synthesis, producing positive-sense progeny HCV-RNA. Newly synthesized HCV-RNAs are utilized for translation and replication, assembling near cytosolic lipid droplets. Assembly involves fusion with luminal lipid droplets in the ER. Subsequent transportation and maturation occur through the Golgi-mediated very-low-density lipoproteins (VLDLs) pathway before packaging and release. In exosomal HCV, the exosome surface reportedly contains CD81, CD63, and HCV E2 protein, while ESCRT-related proteins such as HRS, TSG101, VPS4B, and ALIX are localized within exosomal HCV. Although the entry process for exosomal HCV remains unclear, it may occur through fusogenic or endocytic pathways. Exosomal HCV can be released into cells either by direct fusion with the plasma membrane or through early endosomes from the endocytic pathway, which may fuse with multivesicular bodies (MVBs). MVBs subsequently face two possible fates: fusion with lysosomes for the release of the viral genome or re-secretion into the extracellular space as an exosome via a Rab9A-dependent route after fusion with the plasma membrane. The crosstalk between canonical and exosomal HCV replication and transmission may involve Annexin A2 and ESCRT machineries like HRS, ALIX, TSG101, VPS4B, and syntenin, although detailed mechanisms remain unclear. The question mark denotes an unknown or unclear process and molecular mechanism