The UL3 Protein of Herpes Simplex Virus 1 Is Translated Predominantly from the Second In-Frame Methionine Codon and Is Subject to at Least Two Posttranslational Modifications

摘要

The UL3 open reading frame (ORF) has the coding capacity of 235 codons. The proteins reacting with the anti-UL3 antibody form in denaturing polyacrylamide gel bands with apparent Mrs of 34,000, 35,000, 38,000, 40,000, 41,000, and 42,000 and designated 1 to 6, respectively. Studies on their origins revealed the following. (i) The UL3 proteins forming all six bands were present in lysates of cells infected with wild-type virus and treated with tunicamycin or monensin or in cells infected with the mutant lacking the gene encoding the US3 protein kinase. (ii) The proteins contained in the slower-migrating bands were absent from cells infected with the mutant lacking the UL13 protein kinase. Bands 1 and 3, however were phosphorylated in cells infected with this mutant. (iii) Band 2 protein was absent from cells transfected with a plasmid carrying a substitution of the predicted first methionine codon of the UL3 ORF and superinfected with the UL3⁻ mutant. (iv) Band 1 and 3 proteins were absent from lysates of cells transfected with a plasmid carrying a substitution of the second (M12) methionine codon of the UL3 ORF and superinfected with the UL3⁻ mutant. (v) Cells superinfected with mutants lacking both methionine codons did not accumulate any of the proteins contained in the six bands. (vi) In vitro transcription-translation studies indicated that the translation of band 1 protein was initiated from the second (M12) methionine codon and that band 3 protein represented a UL13-independent, posttranslationally processed form of these proteins. The results indicate that the UL3 protein of herpes simplex virus 1 is translated predominantly from the second in-frame methionine codon and is subject to at least two posttranslational modifications.