Regulation of E-cadherin by mixed lineage kinase 3 (MLK3) mediates cell adhesion in ovarian cancer spheroids

Mixed lineage kinase 3 (MLK3) is a serine/threonine mitogen-activated protein kinase (MAPK) kinase kinase (MAP3K) that activates MAPK signaling pathways and promotes cell proliferation, migration and invasion. E-cadherin is an essential cell adhesion protein that mediates cell-cell interactions in epithelial adherens junctions and is crucial for the integrity of tissues and organs. Here, we demonstrate that MLK3 siRNA knockdown or kinase inhibition significantly impaired spheroid formation in SKOV3, TOV112D and OVCAR3 ovarian cancer cells, that was rescued by overexpression of E-cadherin. Endogenous MLK3 and E-cadherin were co-immunoprecipitated from ovarian cancer cell lysates; and recombinant MLK3 and E-cadherin directly interacted in vitro. MLK3 phosphorylates E-cadherin and has a positive regulatory effect on E-cadherin protein stability. Like E-cadherin, MLK3 is localized to the plasma membrane at regions of cell-cell contact, and MLK3 also has cytoplasmic and nuclear localization. Neither the MLK3-E-cadherin interaction nor the plasma membrane localization of MLK3 is dependent on β-catenin. Collectively, these results indicate a novel function for MLK3 in regulating E-cadherin protein stability, cell adhesion, and ovarian cancer spheroid formation.