Ras adenoviruses modulate cyclin E protein expression and DNA synthesis after partial hepatectomy

Ras-genes encode for proteins important for transmitting extracellular signals from the cytoplasm to the nucleus. In this study we investigated the impact of Ras on cell cycle progression after hepatectomy by using adenoviral vectors (adv) expressing beta-galactosidase (beta-gal), a dominant-negative (Ras N17) or a dominant-active (Ras 61L) form of H-Ras. Partial hepatectomy was performed in mice treated with the different adenoviruses and cell cycle progression was studied by analysing factors involved in cell cycle control during liver regeneration. After hepatectomy, adv Ras 61L increases DNA synthesis significantly in comparison to the other treatment groups. Higher Ras activity results in an early increase of transcriptional active E2F-3, which is associated with higher cyclin E, but almost unchanged cyclin D protein expression. However, Northern blot analysis and cyclin E promoter experiments indicate that, besides transcriptional mechanisms also post-transcriptional mechanisms are involved in regulating cyclin E protein expression after partial hepatectomy in mice treated with adv Ras 61L. Cyclin E phosphorylation studies demonstrate that adv Ras 61L results in hypophosphorylation of cyclin E compared to the control group at early time points after hepatectomy, when cyclin E protein expression strongly increases and there is only a minor effect on cyclin E mRNA levels. Our experiments indicate adv Ras 61L in vivo increases Cyclin E expression by higher transcription via E2F and a post-transcriptional mechanism. These mechanisms result in an earlier activation of an active CDK2/Cyclin E complex which, in turn, triggers DNA synthesis.