Bieri, J.; Suter, C.; Caliaro, O.; Bartetzko, S.; Bircher, C.; Ros, C.

Human parvovirus B19 (B19V), like most parvoviruses, possesses phospholipase A2 (PLA2) activity, which is thought to mediate endosomal escape by membrane disruption. Here, we challenge this model and find evidence for a mechanism of B19V entry mediated by the glycosphingolipid globoside without endosome disruption and retrograde transport to the Golgi. We show that B19V PLA2 activity requires specific calcium levels and pH conditions that are not optimal in endosomes. Accordingly, endosomal membrane integrity was maintained during B19V entry. Furthermore, endosomes remained intact when loaded with MS2 bacteriophage particles pseudotyped with multiple B19V PLA2 subunits, providing superior enzymatic potential compared to native B19V. In globoside knockout cells, incoming viruses are arrested in the endosomal compartment and the infection is blocked. Infection can be rescued by promoting endosomal leakage with polyethyleneimine (PEI), demonstrating the essential role of globoside in facilitating endosomal escape. Incoming virus colocalizes with Golgi markers and interfering with Golgi function blocks infection, suggesting that globoside-mediated entry involves the Golgi compartment, which provides conditions favorable for the lipolytic PLA2. Our study challenges the current model of B19V entry and identifies globoside as an essential intracellular receptor required for endosomal escape.