The synthesis of α (immediate-early) polypeptides in Vero cells infected with pseudorabies virus was studied. Cycloheximide was added at the beginning of infection and removed several hours later. The accumulated α mRNA was translated either in vivo in the presence of actinomycin D to prevent further mRNA synthesis, or in vitro. In intact cells three electrophoretically distinct virus-specific proteins were synthesized, with apparent molecular weights of approximately 180000 (A), 190000 (B) and 200000 (C). The accumulation of B and C was prevented by the proline analogue azetidine. Only protein A was detected in vitro. Proteins B and C were not detected in normally infected cells. All three were associated with the nuclear fraction of cell homogenates and A and B were phosphorylated. The radioactivity of B and C declined during a chase period while that of A increased. This change was prevented by adding cycloheximide during the chase. The pattern of chymotrypsin digestion products suggested that A and B at least were similar proteins. It is presumed that protein A is the single immediate-early protein previously described and analogous to ICP 4 of herpes simplex virus. The significance and function, if any, of proteins B and C is not known but it is possible that they represent stages in the formation or transport of A within the cell and that the progression depends on an unstable protein which is depleted in cells treated with cycloheximide.
Ben-PoratT.,
KervinaM.,
KaplanA. S.1975; Early functions of the genome of herpes virus. V. Serological analysis of ‘immediate-early’ proteins. Virology 65:355–362
ClevelandD. W.,
FischerS. G.,
KirschnerM. W.,
LaemmliU. K.1977; Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. Journal of Biological Chemistry 252:1102–1106
DixonR. A. F.,
SchafferP. A.1980; Fine structure mapping and functional analysis of temperature-sensitive mutants in the gene encoding the HSV-1 immediate-early protein VP175. Journal of Virology 36:189–203
FeldmanL.,
RixonF.,
JeanJ.-H.,
Ben-PoratT.,
KaplanA. S.1979; Transcription of the genome of pseudorabies virus (a herpesvirus) is stringently controlled. Virology 97:316–327
FeldmanL. T.,
DemarchiJ. M.,
Ben-PoratT.,
KaplanA. S.1982; Control of abundance of immediate-early mRNA in herpesvirus (pseudorabies)-infected cells. Virology 116:250–263
FenwickM. L.,
ClarkJ.1982; Expression of early viral genes: a possible pre-α protein infected with HSV. Biochemical and Biophysical Research Communications 108:1454–1459
FenwickM. L.,
ClarkJ.1983; The effect of cycloheximide on the accumulation and stability of functional α-mRNA in cells infected with herpes simplex virus. Journal of General Virology 64:1955–1963
FenwickM. L.,
WalkerM. J.,
PetkevichJ. M.1978; On the association of virus proteins with the nuclei of cells infected with herpes simplex virus. Journal of General Virology 39:519–529
HonessR. W.,
RoizmanB.1974; Regulation of herpes virus macromolecular synthesis. I. Cascade regulation of the synthesis of three groups of viral proteins. Journal of Virology 14:8–19
HonessR. W.,
RoizmanB.1975; Regulation of herpesvirus macromolecular synthesis: sequential transition of polypeptide synthesis requires functional viral polypeptides. Proceedings of the National Academy of Sciences, U. S. A 72:1276–1280
PereiraL.,
WolffM. H.,
FenwickM.,
RoizmanB.1977; Regulation of herpesvirus macromolecular synthesis. V. Properties of α polypeptides made in HSV-1 and HSV-2 infected cells. Virology 77:733–749
PostL. E.,
RoizmanB.1981; A generalized technique for deletion of specific genes in large genomes: α gene 22 of HSV-1 is not essential for growth. Cell 25:227–232
PrestonV. G.1981; Fine-structure mapping of HSV-1 temperature-sensitive mutations within the short repeat region of the genome. Journal of Virology 39:150–161
RakusanovaT.,
Ben-PoratT.,
HimenoM.,
KaplanA. S.1971; Early functions of the genome of herpesvirus. Characterization of the RNA synthesized in cycloheximide-treated infected cells. Virology 46:877–889