A number of genetic events are needed to change the RAS signal from stop to go.
The transforming properties of RAS have been apparent since its discovery 30 years ago. But why does oncogenic RAS cause cell-cycle arrest when introduced into many types of normal cells? Normal cells seem to contain safety mechanisms that inhibit cell-cycle progression when RAS is activated. So, for RAS to have a positive effect on proliferation, additional genetic changes must occur to overcome cell-cycle checkpoint controls and to contribute to the development of tumours. In the April issue of Molecular and Cellular Biology, Alison Lloyd and colleagues show that the p53 and RB pathways have distinct roles in this process.
The authors used primary rat Schwann cells — these proliferate indefinitely, but retain the cell-cycle controls that are normally lost during the immortalization process — to investigate which genetic changes cooperate with oncogenic Ras and contribute to a transformed phenotype. They first investigated the contribution of p53, by expressing oncogenic Ras and either a dominant-negative mutant of p53 (Ras/dnp53) or SV40 large T antigen (Ras/LT), which inhibits the p53 and Rb pathways. Both Ras/dnp53 and Ras/LT cells were able to proliferate in the absence of mitogens, but only Ras/LT cells had a high proliferative capacity in soft agar and within monolayers. So, perhaps both the RB and p53 pathways must be inhibited for RAS to overcome anchorage dependence and contact inhibition.
Deletion analysis of LT showed that the amino terminus — which inhibits the Rb pathway — co-operates with Ras and dnp53 to allow growth in soft agar. Cells expressing dnp53 and activated Ras sometimes spontaneously acquire anchorage independence, with expression of the cyclin-dependent kinase inhibitor Ink4a being lost in 5 out of 10 cases due to promoter methylation. As INK4A lies upstream of the RB family in the tumour-suppressor pathway, its loss would be expected to substitute for the loss of RB-family members and provide the ability to grow independently of anchorage. This was confirmed by showing that antisense Ink4a and expression of a Cdk4 mutant that mimics the loss of Ink4a confer anchorage independence to Ras/dpn53 cells.
So, a number of genetic events are needed to change the RAS signal from stop to go. p53 loss confers mitogen independence, but the RB pathway must also be inhibited for cells to overcome contact inhibition and anchorage dependence. This RB-pathway inhibition frequently occurs through loss of INK4A, which provides an explanation for how this common tumour suppressor contributes to the tumour phenotype.
Emma Croager
References
Mitchell, P. J., Perez-Nadales, E., Malcom, D. S. & Lloyd, A. C. Dissecting the contribution of p16INK4A and the Rb family to the Ras transformed phenotype. Mol. Cell. Biol.23, 2530–2542 (2003) | Article | PubMed |
