Malignant transformation in vivo as well as in vitro requires the activation of oncogenes concomitant with the inactivation of tumor suppressor genes. Although data on the activities and functions of single oncogenes such as c-myc and c-Ha-ras are abundant, less information is available on their synergistic action. We found that primary rat embryo cell lines transformed by c-myc and c-Ha-ras were prone to apoptosis after inhibition of MEK, a downstream kinase of the main Ras signaling pathway. Addition of the MEK1 inhibitor U0126 to a series of myc/ras-expressing cell lines uniformly resulted in unphosphorylated ERK1 and ERK2, morphological alterations and cell cycle arrest, and apoptosis occurred to various extents. The MR-6 cell line apoptosed most promptly after inhibition of MEK1. We analyzed in detail the kinetics of gene regulation of MR-6 cells during apoptosis by means of complementary DNA arrays for the expression of 1,200 characterized rat genes. A twofold increase in expression or reduction of expression below 50% was chosen as the threshold. More than 100 genes were found to be regulated during the early stage of apoptosis induction. One hour after addition of U0126, numerous growth factor receptor genes were induced. By 3 h, G1- or S-phase-specific genes, genes of the AP-1 transcription factor complex and genes coding for cell-surface antigens were upregulated. By 6 h, when MR-6 cells started to apoptose, the expression of most of them returned to baseline values. Data obtained with cDNA arrays were confirmed by northern and western blot analysis. The expression pattern and kinetics of gene regulation in U0126-treated MR-6 cells were compared with those of the MR-2 cell line, in which inhibition of MEK1 resulted in G1 arrest but not cell death. We conclude that myc-mediated apoptosis is accompanied by a transient wave of upregulation of a complex array of genes. At least some of them might represent an abortive attempt by cells to avert cell death.