Characterization of a Herpes Simplex Virus 1 (HSV-1) Chimera in Which the Us3 Protein Kinase Gene Is Replaced with the HSV-2 Us3 Gene.
Us3 protein kinases encoded by herpes simplex virus 1 (HSV-1) and 2 (HSV-2) play important roles in viral replication and pathogenicity. To investigate type-specific differences between HSV-1 Us3 and HSV-2 Us3 in cells infected by viruses with all the same viral gene products except for their Us3 kinases, we constructed and characterized a recombinant HSV-1 in which its Us3 gene was replaced with the HSV-2 Us3 gene. Replacement of HSV-1 Us3 with HSV-2 Us3 had no apparent effect on viral growth in cell cultures or on the range of proteins phosphorylated by Us3. HSV-2 Us3 efficiently compensated for HSV-1 Us3 functions, including blocking apoptosis, controlling infected cell morphology, and downregulating cell surface expression of viral envelope glycoprotein B. In contrast, replacement of HSV-1 Us3 by HSV-2 Us3 changed the phosphorylation status of UL31 and UL34, which are critical viral regulators of nuclear egress. It also caused aberrant localization of these viral proteins and aberrant accumulation of primary enveloped virions in membranous vesicle structures adjacent to the nuclear membrane, and it reduced viral cell-cell spread in cell cultures and pathogenesis in mice. These results clearly demonstrated biological differences between HSV-1 Us3 and HSV-2 Us3, especially in regulation of viral nuclear egress and phosphorylation of viral regulators critical for this process. Our study also suggested that the regulatory role(s) of HSV-1 Us3, which was not carried out by HSV-2 Us3, was important for HSV-1 cell-cell spread and pathogenesis in vivo.


OBJECTIVE
A previous study comparing the phenotypes of HSV-1 and HSV-2 suggested that the HSV-2 Us3 kinase lacked some of the functions of HSV-1 Us3 kinase. The difference between HSV-1 and HSV-2 Us3 kinases appeared to be due to the fact that some Us3 phosphorylation sites in HSV-1 proteins are not conserved in the corresponding HSV-2 proteins. Therefore, we generated recombinant HSV-1 strains YK781 (Us3-chimera) with HSV-2 Us3 and its repaired virus YK783 (Us3-repair) with HSV-1 Us3, to compare the activities of HSV-1 Us3 and HSV-2 Us3 in cells infected by viruses with the same HSV-1 gene products except for their Us3 kinases. We report here that some processes in viral nuclear egress and pathogenesis in vivo that have been attributed to HSV-1 Us3 could not be carried out by HSV-2 Us3. Therefore, our study clarified the biological differences between HSV-1 Us3 and HSV-2 Us3, which may be relevant to viral pathogenesis in vivo.
Division of Molecular Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan Department of Infectious Disease Control, International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Keiko
K
Shindo
Division of Molecular Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan Department of Infectious Disease Control, International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Akihisa
A
Kato
Division of Molecular Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan Department of Infectious Disease Control, International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Naoto
N
Koyanagi
Medical Proteomics Laboratory, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Hiroshi
H
Sagara
Division of Molecular Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan Department of Infectious Disease Control, International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Jun
J
Arii
Division of Molecular Virology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan Department of Infectious Disease Control, International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan ykawagu@ims.u-tokyo.ac.jp.
Yasushi
Y
Kawaguchi
D000818:Animals; D017209:Apoptosis; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000821:virology; D002467:Cell Nucleus / Q000821:virology; D004195:Disease Models, Animal; D005260:Female; D005786:Gene Expression Regulation; D005816:Genetic Complementation Test; D006561:Herpes Simplex / Q000473:pathology / Q000821:virology; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000254:growth & development / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D008813:Mice, Inbred ICR; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D011995:Recombination, Genetic; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D057074:Virus Release; D014779:Virus Replication
D009687:Nuclear Proteins; C108721:UL31 protein, Human herpesvirus 1; C403420:UL34 protein, Human herpesvirus 1; D014764:Viral Proteins; D017346:Protein Serine-Threonine Kinases; C095207:US3 protein, Human herpesvirus 1; C084395:US3 protein, Human herpesvirus 2