Ubiquitin-dependent regulation of phospho-AKT Dynamics by the ubiquitin E3 LIGASE, NEDD4-1, in the insulin-like growth factor-1 response

Background: After activation by phosphorylation, phospho-AKT (pAKT) is translocated to nucleus. Results: Ubiquitination of pAKT by NEDD4-1 is coupled to AKT activation at the plasma membrane by insulin-like growth factor (IGF)-1, which promotes pAKT nuclear trafficking. Conclusion: NEDD4-1 is an E3 ligase for pAKT specifically involved in pAKT nuclear trafficking in IGF-1 signaling. Significance: AKT activation and proper subcellular localization requires specific E3 ligases in a ligand-specific manner. AKTis a critical effector kinase downstream of the PI3K pathway that regulates a plethora of cellular processes including cell growth, death, differentiation, and migration. Mechanisms underlying activated phospho-AKT (pAKT) translocation to its action sites remain unclear. Here we show that NEDD4-1 is a novel E3 ligase that specifically regulates ubiquitin-dependent trafficking of pAKT in insulin-like growth factor (IGF)-1 signaling. NEDD4-1 physically interacts with AKT and promotes HECT domain-dependent ubiquitination of exogenous and endogenous AKT. NEDD4-1 catalyzes K63-type polyubiquitin chain formation on AKT in vitro. Plasma membrane binding is the key step for AKT ubiquitination by NEDD4-1 in vivo. Ubiquitinated pAKT translocates to perinuclear regions, where it is released into the cytoplasm, imported into the nucleus, or coupled with proteasomal degradation. IGF-1 signaling specifically stimulates NEDD4-1-mediated ubiquitination of pAKT, without altering total AKT ubiquitination. A cancer-derived plasma membrane-philic mutant AKT(E17K) is more effectively ubiquitinated by NEDD4-1 and more efficiently trafficked into the nucleus compared with wild typeAKT.This study reveals a novelmechanismby which a specific E3 ligase is required for ubiquitin-dependent control of pAKT dynamics in a ligand-specific manner.