| All herpesvirus particles are composed of four distinct layers: a DNA core, an icosahedral capsid, a proteinaceous layer termed the tegument, and a host-derived lipid envelope. The tegument is composed of at least twenty different viral proteins, many of which have distinct roles during virus egress and virus entry. This thesis focuses on the Us3, VP22, and Us2 tegument proteins of Pseudorabies virus (PRV), an alpha-herpesvirus closely related to the human pathogens herpes simplex virus (HSV) and varicella-zoster virus (VZV).; While characterizing the PRV Us3 serine/threonine kinase, we noted that purified virions from the wild-type PRV Becker strain contained Us3, whereas the attenuated Bartha strain did not. In addition, Bartha also failed to incorporate VP22, though other tegument proteins such as UL48 and beta-actin were packaged efficiently. Despite amino acid differences in Us3 and VP22 between Bartha and Becker, we concluded that the inability of Bartha to package Us3 and VP22 was related to mutations in the Bartha genome outside the Us3 and UL49 (VP22) loci.{09}Furthermore, the Becker, Kaplan, and Bartha PRV strains had different requirements for Us3 in replication and cell-to-cell spread in MDBK cells. Though it is uncertain why Bartha does not incorporate Us3 into the tegument, we hypothesize that this molecule is important for the establishment of/reactivation from latency inside the host.; PRV Us2, a small tegument protein of unknown function, was initially identified by Amanda Clase as having a role in neurovirulence in the chick embryo eye model. To begin to understand Us2 function, we screened a yeast two-hybrid library derived from NIH 3T3 cells to identify cellular proteins that bind Us2. We identified the mitogen-activated protein (MAP) kinases ERK1/ERK2 as Us2 interacting proteins. Us2 co-immunoprecipitated with ERK from infected cells, and altered the localization of ERK in transfected cells. In addition, Us2 inhibited the ability of ERK to phosphorylate nuclear targets, such as Elk-1, by sequestering ERK in the cytoplasm. While the purpose of the Us2/ERK interaction is unclear at present, we hypothesize a role for Us2 in regulation of apoptosis and/or interference with the host innate immune response. |
