Abstract
Abstract
Despite the remarkable success of BCR:ABL1-targeted tyrosine kinase inhibitors, disease progression and therapeutic resistance remain major challenges in chronic myeloid leukemia (CML), highlighting the need to identify additional molecular mechanisms that sustain leukemic survival. In this study, we identify the HECT-type E3 ubiquitin ligase WWP1 as a previously unrecognized regulator of RUNX1 and define a signaling network linking ubiquitination to AMP-activated protein kinase (AMPK)-dependent phosphorylation in CML. We demonstrate that WWP1 directly associates with the PPxY motif of RUNX1 and catalyzes its K63-linked polyubiquitination, thereby enhancing protein stability and promoting leukemic cell growth. Disruption of WWP1, either by genetic silencing or pharmacological inhibition, diminished RUNX1 ubiquitination, impaired proliferation, induced G2/M cell-cycle arrest, and triggered apoptosis. Mechanistic investigations further revealed that WWP1 is a direct substrate of AMPK. Phosphorylation of WWP1 at Thr692 by AMPK promoted its lysosomal degradation, thereby limiting WWP1-dependent ubiquitination of RUNX1. In parallel, AMPK-mediated phosphorylation of RUNX1 reduced its leukemogenic activity, collectively shifting the signaling balance toward growth suppression. Activation of AMPK with metformin reproduced these molecular effects in leukemia cell lines, primary CML patient-derived cells, and xenograft tumors, supporting the physiological relevance of this pathway. Together, our findings establish an AMPK-WWP1-RUNX1 signaling axis that integrates phosphorylation and non-proteolytic ubiquitination to regulate RUNX1 function and CML progression. These results identify WWP1 as a potential therapeutic vulnerability and provide a mechanistic rationale for targeting the AMPK-WWP1 pathway in CML.