FIG. 7.
Coexpression of HPV16 E6 or HPV1 E6 and oncogenic ras in primary human fibroblasts. (A) Determination of p16INK4a levels in POFs coexpressing HPV16 E6 or HPV1 E6 and oncogenic ras. Total protein extracts (200 μg) from cells expressing ras or containing only the empty vector (as indicated) were loaded onto an SDS-15% polyacrylamide gel, transferred onto a PVDF membrane, and incubated with p16INK4a (DCS-50; dilution, 1:250; NovoCastra) or β-tubulin (TUB2.1; dilution, 1:1,000; Sigma) antibody. (B) Growth profiles of POFs coexpressing the viral proteins and oncogenic ras. The growth of the different cell populations was monitored for more than 1 month. Day 0 represents the first day after the completion of antibiotic selection. Differences between the growth curves are statistically significant (pBabe-neo versus HPV16 E6, P < 0.001; pBabe-neo versus HPV1 E6, P < 0.0010). (C) Morphologies of POFs expressing the different viral proteins with oncogenic ras. The different cell populations were photographed on day 30 after selection. Magnifications, ×10.