The relationship between the formation of inclusions and viral DNA synthesis in adenovirus 12-infected cells

Abstract

The relationship between the formation of inclusions and viral DNA synthesis was examined in cells infected with human adenovirus type 12 (H12) DNA-minus temperature-sensitive ( ts) mutants and with H12 wild type (WT) in the presence of inhibitors of DNA synthesis. Three groups of H12 ts mutants ( tsA, tsB, and tsC) are defective in initiation of viral DNA synthesis at the restrictive temperature (40°C). Inclusions were either absent or abortively formed in cells infected with the ts mutants at 40°, although inclusions comparable to those in WT-infected cells were observed in cells infected with the ts mutants at the permissive temperature (34°). The temperature shift-up experiment showed that the inclusion formed in cells infected with tsA at 34° disintegrated at 40°, while those in cells infected with tsB and tsC were retained at 40°. This observation suggests that the morphological integrity of the inclusions is dependent on gene A and the function of the inclusions on genes A, B, and C. The temperature shift-down of tsA-infected cells also showed the formation of inclusions and the recovery in viral DNA synthesis. The inclusions were not formed in WT-infected cells, when viral DNA synthesis was inhibited with either cytosine arabinoside or hydroxyurea. Viral DNA synthesis was not induced and the inclusions were not formed in infected cells after removal of cytosine arabinoside. In contrast, viral DNA synthesis was induced rapidly and the inclusions were formed concomitantly in cells after removal of hydroxyurea. It was shown that the type II inclusions were formed in H12-infected monkey cells, although neither the type IV inclusions nor virions were formed. These observations show the close relationship between the formation of inclusions and viral DNA synthesis, suggesting an important role of DNA-binding protein, the gene-A product, in the formation and the function of inclusions.