SUPPRESSION OF THE PHENOTYPE OF GAMMA(1)34.5(-) HERPES-SIMPLEX VIRUS-1 - FAILURE OF ACTIVATED RNA-DEPENDENT PROTEIN-KINASE TO SHUT OFF PROTEIN-SYNTHESIS IS ASSOCIATED WITH A DELETION IN THE DOMAIN OF THE ALPHA-47 GENE

Earlier studies have shown that infection of human cells by herpes sim plex virus 1 (HSV-1) results in the activation of RNA-dependent protei n kinase (PKR) but that the alpha subunit of eIF-2 is not phosphorylat ed and that protein synthesis is unaffected. In the absence of the vir al gamma(1)34.5 gene, eIF-2 alpha is phosphorylated and protein synthe sis is prematurely shut off (J. Chou, J. J. Chen, M. Gross, and B. Roi zman, Proc. Natl. Acad. Sci. USA 92:10516-10520, 1995), A second recen t paper reported the selection of second-site suppressor mutants chara cterized by near-wild-type protein synthesis in cells infected with ga mma(1)34.5(-) mutants (I. Mohr and Y. Gluzman, EMBO J. 15:4759-1766, 1 996). Here, we report the properties of the spontaneous HSV-1 suppress or mutant Sup-1, which is characterized by spontaneous deletion of 503 bp encompassing the domain of the alpha 47 gene and junction with the inverted repeats flanking the unique short (U-s) sequence of the HSV- 1 DNA resulting in the juxtaposition of the alpha 47 promoter to the c oding domain of the U(s)11 gene. This mutant does not exhibit the shut off of protein synthesis characteristic of the gamma(1)34.5(-) virus. Specifically, Sup-1 in SK-N-SH human neuroblastoma cells (i) did not e xhibit the function of the alpha 47 gene characterized by a reduction in the transport of peptides across the endoplasmic reticulum of perme alized cells consistent with the absence of alpha 47 gene sequences, ( ii) accumulated U(s)11 protein at levels analogous to those of the wil d-type parent but the protein was made at earlier times after infectio n, as would be expected from a change in the promoter, and (iii) activ ated PKR like that of the parent, gamma(1)34.5(-) virus, but (iv) did not cause premature shutoff of protein synthesis and therefore was sim ilar to the wild-type parent virus rather than the gamma(1)34.5(-) vir us from which it was derived, We conclude that the mechanism by which Sup-1 blocks the shutoff of protein synthesis associated with phosphor ylation of eIF-2 alpha by the activated PKR is not readily explainable by a secondary mutation characterized by a deletion.