Expression of Rap 1 suppresses genomic instability of H-ras transformed mouse fibroblasts.

Among the multiple genetic changes that occur during cancer progression are the activation of proto-oncogenes and the inactivation or loss of genes encoding tumor suppressors. The potential roles for these genes in the perturbation of genome stability continues to be of major interest. We have previously shown that conditional expression of H-ras in NIH3T3 cells increases genetic instability in these cells, rendering them more permissive to gene amplification and to the generation of chromosome aberrations which can be induced within a single cell cycle. In the present study we show that genetic instability induced by H-ras expression can be suppressed by co-expression of Rap 1, a Ras-related tumor suppressor gene. An NIH3T3 cell line transformed with activated human H-ras was transfected with Rap 1. Expression of the Rap 1 gene reverted the transformed cells to a flat morphology. The reverted cells reestablished contact inhibition of growth and lost the capacity to form colonies in soft agar. These cells were subsequently studied for the role of Rap 1 on the suppression of genomic instability induced by oncogenic H-ras. Cells transformed with H-ras manifest an increase in methotrexate resistance as measured by an increase in Dhfr gene amplification. Cells which concommitantly express Rap 1 showed reduced levels of methotrexate resistance as well as reduction of gene amplification capacity. Furthermore fluorescent-in-situ hybridization (FISH) with a pancentromeric mouse probe showed that elevated levels of chromosome aberrations in cells expressing H-ras were also suppressed after co-expression of Rap 1.