Cdc14 is a member of the dual-specificity phosphatase family, which is essential for faithful cell cycle progression in eukaryotic cells of different origin. The function of human Cdc14A (hCdc14A), however, has not been fully elucidated as only few physiological substrates have been identified. To gain insight into the biological role of Cdc14A, we performed a yeast two-hybrid screen designed to isolate substrates of this human phosphatase. Using this genetic approach, we here report the identification of Erk3, an atypical mitogen-activated protein kinase (MAPK), as a specific binding partner of hCdc14A. GST pull-down assays show that Erk3 interacts directly with hCdc14A in vitro via its unique C-terminal domain. Furthermore, biochemical analysis reveals that hCdc14A can remove cyclin-dependent kinase (Cdk)-mediated phosphorylation of Erk3 in vitro raising the possibility that Erk3 may be a potential substrate for hCdc14A in vivo. Consistent with a physiologically relevant cross-talk in vivo, we find that Cdc14A forms a stable complex with Erk3 in human cells independent of its intrinsic phosphatase activity but mediated by its regulatory C-terminal domain. We show that hCdc14A impacts the emerging signaling pathway between Erk3 and MK5, a MAPK-activated protein kinase. We document that hCdc14A upregulation leads to redistribution of the Erk3 substrate MK5 from the nucleus to the cytoplasm. In addition, we find that hCdc14A stabilizes complex formation between Erk3 and its binding partner cyclin D3, a D-type cyclin implicated in both cellular proliferation and differentiation. Collectively, our findings suggest an intimate functional relationship between the Cdc14A phosphatase and the Erk3 kinase in signaling pathways that regulate key cell-fate decisions in human cells.

Institute of Cancer Biology and Centre for Genotoxic Stress Research Danish Cancer Society Copenhagen Denmark