Ullah, K.

Hypoxia and NF-kB signaling are well-established drivers of cancer progression and treatment failure, yet the oxygen-dependent regulation of non-canonical NF-kB signaling remains poorly defined. Here, we identify hypoxia-inducible factor prolyl-4-hydroxylase-1 (HIF-P4H-1/EGLN2) as a key modulator of the non-canonical NF-kB pathway. Using integrated biochemical, genetic, proteomic, and transcriptomic analyses across human cell lines, mouse models, and clinical tumor samples, we demonstrate that HIF-P4H-1 directly interacts with and hydroxylates IKKa at proline 367, thereby promoting its ubiquitination and proteasomal degradation. Loss or inhibition of HIF-P4H-1 results in accumulation of IKKa, impaired NF-kB2/p100 processing to p52, destabilization of NF-kB-inducing kinase (NIK), and suppression of non-canonical NF-kB-dependent survival gene expression. Structural modeling and mutagenesis identify proline 367 hydroxylation as a critical determinant of IKKa turnover. Analysis of TCGA cohorts reveals an inverse correlation between HIF-P4H-1 and IKKa expression, with elevated HIF-P4H-1 associating with advanced tumor stage and reduced overall survival in clear cell renal cell carcinoma. Functionally, targeting HIF-P4H-1 sensitizes cancer cells to cell death and impairs proliferation, clonogenic growth, and migration. Together, our findings define a previously unrecognized oxygen-dependent mechanism regulating non-canonical NF-kB signaling through direct control of IKKa stability and nominate the HIF-P4H-1-IKKa axis as a potential therapeutic vulnerability in hypoxia-adapted malignancies.