Differences in the intracellular localization and fate of herpes simplex virus tegument proteins early in the infection of Vero cells.
The fate of herpes simplex virus 1 (HSV-1) tegument proteins during infection in Vero cells was investigated immunochemically. Input virion-associated VP13/14 and VP16 localized to the nucleus early in infection, while VP1/2 localized to the nuclear envelope of the cell and VP22 could not be detected using monoclonal antibody P43. Western blotting suggested that virion-associated VP13/14, VP16 and VP22 were stable in infected cells whereas VP1/2 appeared to be processed or modified. Further studies showed that P43 recognized a phosphorylation-sensitive epitope in VP22 and suggested that virion-associated VP22 was phosphorylated upon entry to the cell. VP13/14 and VP16 were easily extracted from cells early in infection whereas VP22 was largely insoluble. Phosphatase treatment of soluble extracts caused a shift in the molecular mass of VP16 showing it was phosphorylated. As infection progressed VP16 was observed in discrete nuclear compartments where it co-localized with ICP8 and the capsid-associated protein VP22a. VP13/14 was also observed in the nucleus. P43 immunostaining appeared around 6 h post-infection as punctate nuclear foci which often localized to the edge of VP16-immunoreactive areas. Punctate P43 cytoplasmic staining appeared around 12 h post-infection. By 18 h the nuclear pattern had disappeared and an extensive cytoplasmic stain was observed which closely overlapped that of other tegument proteins. On the basis of these data we suggest that virion-associated VP22 is phosphorylated upon entry of the virus into the cell and that unphosphorylated VP22, which is preferentially recognized by P43, becomes available later in infection, initially in the nucleus, for packaging into virions.
Molecular Medicine Unit, University of Leeds, St James's University Hospital, UK.
E E
EE
Morrison
A J
AJ
Stevenson
Y F
YF
Wang
D M
DM
Meredith
D000818:Animals; D015153:Blotting, Western; D002467:Cell Nucleus / Q000737:chemistry; D002522:Chlorocebus aethiops; D003513:Cycloheximide / Q000494:pharmacology; D018259:Herpesvirus 1, Human / Q000737:chemistry*; D010766:Phosphorylation; D014709:Vero Cells; D014764:Viral Proteins / Q000032:analysis*; D014771:Virion / Q000737:chemistry*
D014764:Viral Proteins; D003513:Cycloheximide