Mixed Lineage Kinase 3 Modulates β-Catenin Signaling in Cancer Cells

Expression of beta-catenin is strictly regulated in normal cells via the glycogen synthase kinase 3 beta (GSK3 beta)-adenomatous polyposis coli-axin-mediated degradation pathway. Mechanisms leading to inactivation of this pathway (example: activation of Wnt/beta-catenin signaling or mutations of members of the degradation complex) can result in beta-catenin stabilization and activation of beta-catenin/T-cell factor (TCF) signaling. beta-Catenin-mediated cellular events are diverse and complex. A better understanding of the cellular signaling networks that control beta-catenin pathway is important for designing effective therapeutic strategies targeting this axis. To gain more insight, we focused on determining any possible cross-talk between beta-catenin and mixed lineage kinase 3 (MLK3), a MAPK kinase kinase member. Our studies indicated that MLK3 can induce beta-catenin expression via post-translational stabilization in various cancer cells, including prostate cancer. This function of MLK3 was dependent on its kinase activity. MLK3 can interact with beta-catenin and phosphorylate it in vitro. Overexpression of GSK3 beta-WT or the S9A mutant was unable to antagonize MLK3-induced stabilization, suggesting this to be independent of GSK3 beta pathway. Surprisingly, despite stabilizing beta-catenin, MLK3 inhibited TCF transcriptional activity in the presence of both WT and S37A beta-catenin. These resulted in reduced expression of beta-catenin/TCF downstream targets Survivin and myc. Immunoprecipitation studies indicated that MLK3 did not decrease beta-catenin/TCF interaction but promoted interaction between beta-catenin and KLF4, a known repressor of beta-catenin/TCF transcriptional activity. In addition, co-expression of MLK3 and beta-catenin resulted in significant G(2)/M arrest. These studies provide a novel insight toward the regulation of beta-catenin pathway, which can be targeted to control cancer cell proliferation, particularly those with aberrant activation of beta-catenin signaling.