HGS+ enlarged vesicular compartment serves as site for coronavirus assembly at later infection stage

The precise site of coronavirus assembly remains poorly characterized. In this study, we observed that coronavirus infection induces the host factor hepatocyte growth factor-regulated tyrosine kinase substrate (HGS) to form distinct HGS enlarged vesicular compartments at later infection stage. Confocal and live-cell super-resolution microscopy showed that viral structural proteins colocalize with these HGS enlarged vesicular compartments. Correspondingly, APEX2-based electron microscopy (APEX2-EM) and immuno-EM analyses confirmed the presence of assembled virions within these unique HGS compartments, identifying them as sites of virion assembly. This was further supported by cryogenic correlated light and electron microscopy (cryo-CLEM), which captured ongoing virion formation occurring within HGS enlarged vesicles. Crucially, whole cell volume EM revealed that HGS deficiency abolishes these vesicular compartments and markedly reduces assembled virions. Lastly, we demonstrated that HGS vesicular compartments are rearranged from Golgi and endosome/lysosome by coronavirus infection. Together, these findings establish that coronavirus-induced HGS vesicular compartments function as essential platforms for virion assembly at later infection stage.