In vitro translation of adenovirus type 12-specific mRNA isolated from infected and transformed cells

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Abstract
The early and late gene products of human adenovirus type 12 (Ad12) and the viral proteins synthesized in an Ad12-transformed cell line were identified by translation of viral mRNA in an in vitro protein-synthesizing system. Cytoplasmic RNA was isolated from permissive KB [human oral carcinoma] or nonpermissive BHK [baby hamster kidney] cells infected with Ad12 and from Ad12-transformed HA12/7 [Syrian hamster kidney] cells. Virus-specific RNA was selected by hybridization to Ad12 DNA covalently bound to cellulose. Viral RNA was then translated in a fractionated rabbit reticulocyte cell-free system or in wheat germ S-30 extracts. The proteins synthesized were characterized by immunoprecipitation and subsequent electrophoresis on sodium dodecyl sulfate-polyacrylamide gels. RNA prepared from KB cells late after infection with Ad12 elicited the synthesis of most of the structural polypeptides of the virion and at least 2 presumably nonstructural Ad12 proteins. When viral RNA isolated early after infection of KB cells with Ad12 was translated in vitro, 10 polypeptides were observed: E-68K, E-50K, E-42K, E-39K, E-34K, E-21K, E-19K, E-13K, E-12K and E-10K. Ad12-specific RNA was also isolated from the Ad12-transformed hamster cell line HA12/7, which contains several copies of the Ad12 genome integrated in the host genome. The RNA codes for at least 7 polypeptides with MW very similar to those of the early viral proteins.