Prolyl-4-Hydroxylase Domain (PHD) enzyme 2 is one of three PHD isoforms that regulate HIF-1a stabilization by hydroxylation of HIF-1a prolines. Stabilized HIF-1 confers the expression of a set of oxygen-dependent genes. Analysis of PHD2 knockdown should deliver insight into PHD2-dependent cell processes. Therefore we analysed the tetracycline-inducible PHD2 knockdown T-REx HeLa cell line P2.16.

Transcriptome and proteome changes revealed genes and proteins as regulated that play a role within cell locomotion and cell spreading processes. We found a PHD2-dependent gene expression of tetraspanin 8 (TSPAN8, TM4SF3, CO-029) – a gene encoding for a transmembrane protein that might be involved in cancer progression and metastasis – as well as a PHD2-dependent phosphorylation of the actin depolymerizing factor Cofilin. The impact of PHD2 on these genes and proteins correlated with a phenotypical change of tetracycline treated P2.16 compared with non treated cells regarding cell motility and adhesion.

Further we investigated the molecular involvement of the RhoA/ROCK pathway in PHD2-dependent phosphorylation of cofilin, since this pathway is known to regulate cofilin activity and thereby cytoskeletal reorganization.