Induction of cellular DNA synthesis by adenovirus type 12 in a set of temperature-sensitive mutants of rat 3Y1 fibroblasts blocked in G1 phase

Four temperature-sensitive (ts) mutants of rat 3Y1 fibroblasts, which represent separate complementation groups, cease to proliferate predominantly with a 2C DNA content, either at 39.8 degrees (temperature arrest), or at 33.8 degrees at a confluent cell density (density arrest). When infected at 39.8 degrees with adenovirus type 12 (Ad12), cells of all four ts mutants in both arrest states entered the S phase, thereby suggesting that Ad12 overcomes the four independent functional blocks to cellular entry into S phase. Results of experiments using Ad12 E1-region mutants suggest that the E1A gene product(s) is indispensable to overcoming the ts block, whereas the E1B product(s) may be dispensable. The cell killing observed in 3Y1 cells infected with wild-type Ad12 did not occur in infection with one of the E1-region mutants with a 6-bp insertion in the E1A 13 S mRNA unique region. When infected with this mutant at 39.8 degrees, two ts mutants of 3Y1 (3Y1tsF121 and 3Y1tsG125) in both arrested states proliferated through at least one generation. Another mutant (3Y1tsD123) was accelerated to die following entry into the S phase. In the other mutant (3Y1tsH203), the cell number was either unchanged (temperature arrest) or was increased less than twofold and then decreased (density arrest). The findings with the latter two mutant lines suggest that induction of cellular DNA synthesis is not sufficient for the subsequent proliferation of the infected cells, and that the Ad12 gene function(s) does not directly rescue the primary lesions in these ts mutants but does overcome some of the blocks to concomitantly occurring events. In the former two mutant lines, however, Ad12 gene function(s) may directly rescue the ts lesions. We propose that the Ad12 gene product(s) can overcome blocks to the initiation of cellular DNA synthesis but cannot overcome blocks to events related to cell survival.