The cytoplasmic, poly(A)-containing RNA species complementary to all regions of the adenovirus type 2 (Ad2) genome expressed before the onset of viral DNA synthesis have been examined by "Northern" blotting. HeLa cells infected with low multiplicities of Ad2 were harvested at 2-hr intervals until late in the infectious cycle to establish the temporal patterns of expression of each viral gene. Under these conditions, such late mRNA products as those of the L3 and L5 families were not detected until 14 hr after infection. Although individual mRNA species complementary to several genes showed different patterns of expression, the order of appearance in the cytoplasm of substantial concentrations of adenoviral mRNA species was E1A, E3, and E4 (4 to 6 hr), E2A and E2B (8 hr), 3.7- and 4.1-kb L1 mRNA species (10-12 hr), IX and IVa2 mRNAs (12 hr), and those encoded in the major late transcriptional unit, such as members of the L3 and L5 families (14 hr). The mRNA species encoding polypeptides IX and IVa2 were not produced when viral DNA synthesis was blocked, whereas the larger L1 mRNA species was made under these conditions. Two E2B mRNA species, some 5.0 and 7 kb, were observed at low concentrations at 8 hr after infection and their concentration increased until 24 hr after infection, as did that of the E2A mRNA species: the products of the E2 transcription unit appeared to be expressed coordinately and at a constant ratio throughout infection. Few of the early mRNA species decreased in concentration after the onset of the late phase of infection. Examination of the viral mRNA produced when protein synthesis was inhibited by 10 μM anisomycin added 3 hr after infection suggested that processing of certain viral, early transcripts was altered in the presence of the drug.
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