Prolyl hydroxylase-1 negatively regulates IκB kinase-β, giving insight into hypoxia-induced NFκB activity

Hypoxia is a feature of the microenvironment of a growing tumor. The transcription factor NF kappa B is activated in hypoxia, an event that has significant implications for tumor progression. Here, we demonstrate that hypoxia activates NFKB through a pathway involving activation of I kappa B kinase-beta (IKK beta) leading to phosphorylation-dependent degradation of I kappa B alpha and liberation of NF kappa B. Furthermore, through increasing the pool and/or activation potential of IKK beta, hypoxia amplifies cellular sensitivity to stimulation with TNF alpha. Within its activation loop, IKK beta contains an evolutionarily conserved LxxLAP consensus motif for hydroxylation by prolyl hydroxylases (PHDs). Mimicking hypoxia by treatment of cells with siRNA against PHD-1 or PHD-2 or the pan-prolyl hydroxylase inhibitor DMOG results in NF kappa B activation. Conversely, overexpression of PHD-1 decreases cytokine-stimulated NF kappa B reporter activity, further suggesting a repressive role for PHD-1 in controlling the activity of NF kappa B. Hypoxia increases both the expression and activity of IKK beta, and site-directed mutagenesis of the proline residue (P191A) of the putative IKK beta hydroxylation site results in a loss of hypoxic inducibility. Thus, we hypothesize that hypoxia releases repression of NF kappa B activity through decreased PHD-dependent hydroxylation of IKK beta, an event that may contribute to tumor development and progression through amplification of tumorigenic signaling pathways.