Kaposi sarcoma-associated herpesvirus gB dictates a low-pH endocytotic entry

All herpesviruses encode a so-called fusion protein, gB, which fuses the virus membrane with host membranes. Fusion is essential for infection by enveloped viruses. Under conditions where gBs from other herpesviruses fuse cells into syncytia when expressed, KSHV gB does not readily fuse cells, and KSHV infection was previously shown to be inhibited by substances that raise the pH of endocytotic vesicles. We therefore sought to test whether fusion of KSHV viral particles with cellular membranes is a pH-dependent step. We developed two tools to test this hypothesis. The first tool is a dual-fluorescent KSHV with differently colored protein tags at the viral envelope and the capsid, which lose colocalization upon fusion and release of the capsid from the membrane, but not in the presence of an inhibitor of vesicle acidification. The second tool is a hybrid RRV strain that expresses KSHV gB instead of RRV gB. While RRV infected receptor-overexpressing cells in a manner that was not inhibited by an inhibitor of vesicle acidification, the hybrid RRV expressing KSHV gB was sensitive to inhibition of vesicle acidification. Together, these findings show that KSHV gB dictates a low-pH, endocytic route of infection.