Role of reactive oxygen species in the regulation of HIF-1 by prolyl hydroxylase 2 under mild hypoxia

The function and survival of eukaryotic cells depends on a constant and sufficient oxygen supply. Cells recognize and respond to hypoxia by accumulation of the transcription factor hypoxia-inducible factor 1 (HIF-1), composed of an oxygen-sensitive HIF-1 alpha and a constitutive HIF-1 beta subunit. Besides physiology, HIF-1 induction is involved in major pathological processes such as cardiovascular disease, inflammation and cancer, which are associated with the formation of reactive oxygen species (ROS). ROS have been reported to affect HIF-1 activity but the role for ROS in regulating HIF-1 has not been definitely settled. In order to shed light on the redox-regulation of HIF-1 by ROS, we studied the impact of exogenous ROS treatment (H2O2) on HIF-1 alpha and HIF-1 regulatory protein prolyl hydroxylase 2 (PHD2) in the human osteosarcoma cell line U2OS. At early reaction periods, H2O2 induced HIF-1 alpha but at prolonged observation phases the opposite occurred. Herein, modulation of PHD activity appeared to be the key element, because knockdown and inhibition of the PHD2 prevented reduction of HIF-1 alpha. However, H2O2 treatment constantly suppressed HIF-1 transactivation at all time-points. Our data indicate a dual redox regulation of HIF-1 alpha protein amount with a constant suppression of HIF-1 target gene expression by ROS.