Functional Domains of the Herpes Simplex Virus Type 1 Tegument Protein pUL37: The Amino Terminus is Dispensable for Virus Replication in Tissue Culture.

The herpes simplex virus type 1 (HSV-1) UL37 gene encodes for a multifunctional component of the virion tegument, which is necessary for secondary envelopment in the cytoplasm of infected cells, for motility of the viral particle, and for the first steps in the initiation of virus infection. This 120 kDa protein has several known viral interacting partners, including pUL36, gK/pUL20, pUS10, and VP26, and cellular interacting proteins which include TRAF6, RIG-I, and dystonin. These interactions are likely important for the functions of pUL37 at both early and late stages of infection. We employed a genetic approach to determine essential domains and amino acid residues of pUL37 and their associated functions in cellular localization and virion morphogenesis. Using marker-rescue/marker-transfer methods, we generated a library of GFP-tagged pUL37 mutations in the HSV-1 strain KOS genome. Through viral growth and ultra-structural analysis, we discovered that the C-terminus is essential for replication. The N-terminal 480 amino acids are dispensable for replication in cell culture, although serve some non-essential function as viral titers are reduced in the presence of this truncation. Furthermore, the C-terminal 133 amino acids are important in so much that their absence leads to a lethal phenotype. We further probed the carboxy terminal half of pUL37 by alanine scanning mutagenesis of conserved residues among alphaherpesviruses. Mutant viruses were screened for the inability to form plaques-or greatly reduced plaque size-on Vero cells, of which 22 mutations were chosen for additional analysis. Viruses discovered to have the greatest reduction in viral titers on Vero cells were examined by electron microscopy (EM) and by confocal light microscopy for pUL37-EGFP cellular localization. This genetic approach identified both essential and non-essential domains and residues of the HSV-1 UL37 gene product. The mutations identified in this study are recognized as significant candidates for further analysis of the pUL37 function and may unveil previously undiscovered roles and interactions of this essential tegument gene.