The DNA binding domain of herpes simplex virus type 1 origin binding protein is a transdominant inhibitor of virus replication.
The origin binding protein (OBP) of herpes simplex virus (HSV) type 1 specifically interacts with two high-affinity sites in each HSV DNA replication origin. The sequence-specific DNA binding activity of OBP maps to the carboxy-terminal one-third of the protein. For a single binding site, recombinantly expressed forms of this DNA binding domain have the same sequence specificity and binding affinity as the full-length OBP. However, unlike the full-length protein, truncated OBP does not bind HSV replication origins in a cooperative manner. To determine if cooperative interactions between DNA-bound OBP molecules are essential for viral DNA replication, the 317-amino-acid carboxy-terminal DNA binding domain of OBP was expressed in chick embryo fibroblasts. Cells were infected with HSV type 1, and viral DNA synthesis and virus production were monitored. We found that cells expressing truncated OBP were severely restricted for virus replication and that HSV DNA synthesis was undetectable. The results demonstrate that the amino-terminal two-thirds of OBP is essential for HSV DNA replication and that the OBP DNA binding domain acts as a transdominant inhibitor of viral DNA replication. The results also suggest that this experimental approach could be used to generate a refined map of essential OBP functions and that the approach may be generally applicable to the analysis of the multifunction HSV DNA replication complex.
Perry
H C
HC
Department of Virus and Cell Biology, Merck Research Laboratories, West Point, Pennsylvania 19486.
Hazuda
D J
DJ
McClements
W L
WL
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002642:Chick Embryo; D004261:DNA Replication / Q000235:genetics / Q000502:physiology; D004279:DNA, Viral / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D005347:Fibroblasts; D008969:Molecular Sequence Data; D018139:Simplexvirus / Q000235:genetics / Q000502:physiology*; D014764:Viral Proteins / Q000235:genetics / Q000502:physiology*; D014779:Virus Replication / Q000235:genetics / Q000502:physiology*
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D014764:Viral Proteins; C064565:UL9 protein, Human herpesvirus 1