The UL12.5 gene product of herpes simplex virus type 1 exhibits nuclease and strand exchange activities but does not localize to the nucleus.
The herpes simplex virus type 1 (HSV-1) alkaline nuclease, encoded by the UL12 gene, plays an important role in HSV-1 replication, as a null mutant of UL12 displays a severe growth defect. Although the precise in vivo role of UL12 has not yet been determined, several in vitro activities have been identified for the protein, including endo- and exonuclease activities, interaction with the HSV-1 single-stranded DNA binding protein ICP8, and an ability to promote strand exchange in conjunction with ICP8. In this study, we examined a naturally occurring N-terminally truncated version of UL12 called UL12.5. Previous studies showing that UL12.5 exhibits nuclease activity but is unable to complement a UL12 null virus posed a dilemma and suggested that UL12.5 may lack a critical activity possessed by the full-length protein, UL12. We constructed a recombinant baculovirus capable of expressing UL12.5 and purified soluble UL12.5 from infected insect cells. The purified UL12.5 exhibited both endo- and exonuclease activities but was less active than UL12. Like UL12, UL12.5 could mediate strand exchange with ICP8 and could also be coimmunoprecipitated with ICP8. The primary difference between the two proteins was in their intracellular localization, with UL12 localizing to the nucleus and UL12.5 remaining in the cytoplasm. We mapped a nuclear localization signal to the N terminus of UL12, the domain absent from UL12.5. In addition, when UL12.5 was overexpressed so that some of the enzyme leaked into the nucleus, it was able to partially complement the UL12 null mutant.
Reuven
Nina Bacher
NB
Department of Molecular, Microbial, and Structural Biology, University of Connecticut Health Center, Farmington, Connecticut 06030-3205, USA.
Antoku
Susumu
S
Weller
Sandra K
SK
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D018259:Herpesvirus 1, Human / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D045522:Recombinases / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism*; D018411:Spodoptera; D014709:Vero Cells; D014764:Viral Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D004268:DNA-Binding Proteins; C045462:ICP8 protein, Simplexvirus; D045522:Recombinases; D014764:Viral Proteins; D012260:Ribonucleases