The Transmembrane and Cytosolic Domains of Equine Herpesvirus Type 1 Glycoprotein D Determine Golgi Retention by Regulating Vesicle Formation

Accumulating evidence suggests that envelope proteins play an important role in viral secondary envelopment; however, the molecular mechanisms involved are poorly understood. To clarify these mechanisms, we studied the localization of equine herpesvirus type 1 (EHV-1) envelope proteins and found that glycoprotein D of EHV-1 (gDEHV−1) was mostly retained in the Golgi complex, unlike that of HSV-1 and PRV. We used a gene truncation and replacement strategy to investigate the determinant sequence responsible for the Golgi retention phenotype and found that Golgi retention signals exhibit multi-domain features. The extracellular domain of gDEHV−1 (ECDEHV−1) is an endoplasmic reticulum (ER)-resident domain. The transmembrane domain and cytoplasmic tail (TM-CT) of gDEHV−1 was found to help the protein reside in the Golgi complex. Once each of the dual domains was deleted or replaced, the mutant gD remained in the ER. (TM-CT)EHV−1 preferentially binds to the endomembrane and induces a large number of vesicles that may originate from the Golgi complex or ER-Golgi intermediate compartment. Membrane fusion was hardly observed between the cell membrane and the induced vesicles. These findings provide further insight into the molecular mechanism underlying the Golgi retention of gDEHV−1, enhancing our understanding of viral secondary envelopment.