The herpes simplex virus type 1 (HSV-1) tegument protein VP16 is a potent transcriptional inducer of immediate-early gene expression, comprising an N-terminal domain involved in binding DNA linked to an acidic transactivating C-terminal domain. The gene encoding the counterpart of this protein in equid herpesvirus 4 (EHV-4) was sequenced. Comparisons with VP16 and the homologous proteins of equine herpesvirus 1 (EHV-1) and varicella-zoster virus (VZV) showed that a region in the N-terminal domain involved in formation of a complex with cellular proteins is partially conserved in all four proteins. In contrast, the C-terminal regions of the EHV proteins, like that of VZV, are not particularly acidic and are not significantly conserved with respect to the C-terminal region of VP16. Nevertheless, transient expression experiments indicated that the EHV-1 and EHV-4 proteins are able to transactivate HSV-1 and EHV-1 immediate-early promoters in a dose-dependent manner, which suggests that this activity is not dependent on an acidic C-terminal domain.