Tumor hypoxia is correlated with genetic alteration and malignant progression. Our previous studies indicated that the hypoxia-inducible transcription factor, HIF-1α, is responsible for hypoxic suppression of DNA repair in tumor cells by a non-canonical mode of action that requires the HIF-1α PAS-B subdomain. The involvement of HIF-1α in genetic alteration has raised an intriguing question as to whether normal cells would respond to hypoxic stress differently to avert genetic alteration. In this study, we chose several mouse cell types ranging from benign to malignant, apoptosis-proficient to apoptosis-deficient, and determined their responses to HIF-1α expression. In agreement with our previous findings, transient hypoxia and HIF-1α expression inhibited DNA repair and induced DNA damage in all cell types examined; however, cumulative DNA damage only occurred in apoptosis-deficient, malignant cells transduced for sustained expression of HIF-1α or HIF-1α PAS-B itself. In keeping with the theory of apoptosis as a cancer barrier, only these apoptosis-deficient cells acquired anchorage-independent growth and epithelial-mesenchymal transition. Furthermore, these cells exhibited increased Akt activity and resistance to etoposide by inhibiting autophagy. Altogether, our results define an essential role for apoptosis to prevent HIF-1α-induced genetic alteration and thereby malignant progression.