Role of the cytoplasmic tails of pseudorabies virus glycoproteins B, E andM in intracellular localization and virion incorporation

The cytoplasmic domains of several herpesviral glycoproteins encompass pote ntial intracellular sorting signals. To analyse the function of the cytopla smic domains of different pseudorabies virus (PrV) glycoproteins, hybrid pr oteins were constructed consisting of the extracellular and transmembrane d omains of envelope glycoprotein D (gD) fused to the cytoplasmic tails of gB , gE or gM (designated gDB, gDE and gDM), all of which contain putative end ocytosis motifs, gD is a type I membrane protein required for binding to an d entry into target cells. Localization of hybrid proteins compared to full -length gB, gE and gM as well as carboxy-terminally truncated variants of g D was studied by confocal laser scanning microscopy. The function of gD hyb rids was assayed by trans-complementation of a gD-negative PrV mutant. The carboxy-terminal domains of gB and gM directed a predominantly intracellula r localization of gDB and gDM, while full-length gD and a tailless gD mutan t (gDc) were preferentially expressed on the cell surface. In contrast gDE, and a gDB lacking the putative gB endocytosis signal (gDB Delta 29), were predominantly located in the plasma membrane. Despite the different intrace llular localization, all tested proteins were able to complement infectivit y of a PrV gD(-) mutant. Cells which stably express full-length gD and plas ma-membrane-associated gD hybrids exhibit a significant resistance to PrV i nfection, while cells expressing predominantly intracellularly located form s do not. This suggests that the assumed sequestration of receptors by gD, which is supposed to be responsible for the interference phenomenon, occurs at the cell surface.