| Introduction: Plant homeodomain protein Jade-1, which is stabilized by the von Hippel-Lindau tumor suppressor, is a candidate renal tumor suppressor that functions as a ubiquitin ligase to inhibit canonical Wnt signaling and also as a transcription factor associated with histone acetyltransferase activity.;Hypothesis: Jade-1 may be involved in additional signaling pathways to function as a renal tumor suppressor, as Jade-1 homolog in Drosophila can antagonize Ras signaling.;Results: Kinase arrays were used to identify additional signaling pathways in which Jade-1 may be involved. Jade-1 knockdown increased levels of endogenous phospho-AKT1 (pAKT1), and overexpression of Jade-1 decreased levels of endogenous pAKT1. Likewise, in HK-2 cells, a differentiated renal proximal tubular cell line and a model of renal cancer precursor cells, levels of endogenous pAKT1 and pAKT were considerably increased with Jade-1 silencing and inhibited by reintroduction of Jade-1 or by specific PI3K inhibitors. Examination of pAKT downstream targets and in vitro AKT kinase assays confirmed the effect of Jade-1 on AKT activity. Jade-1-silenced HK-2 cell lines grew faster than non-silenced control cells. Intriguingly, Jade-1-silenced HK-2 cells acquired AKT-dependent anchorage-independent growth, which was inhibited by reintroduction of Jade-1, PI3K inhibition, or dominant-negative AKT and aggravated by constitutively active AKT. Importantly, constitutively active AKT alone could not fully substitute for the effect of Jade-1-silencing on anchorage-independent growth, suggesting deregulation of additional signaling pathways in Jade-1-silenced cells.;Jade-1 also negatively regulates Wnt signaling by ubiquitinating beta-catenin. We therefore treated multiple renal cancer cell lines with small-molecule inhibitors of beta-catenin, hexachlorophene or PKF115-584 (Novartis). Growth of renal cancer cells was greatly inhibited with an IC50 of around 10 muM for hexachlorophene and 0.3 muM for PKF115-584. Expression of cyclin D1 and c-Myc, two Wnt target genes, was greatly downregulated by PKF115-584. Anchorage-independent growth of Jade-1-silenced HK-2 cells was also abolished by PKF115-584.;Conclusion: Existing evidence indicates that multiple points of crosstalk exist between the PI3K-AKT and the beta-catenin pathway, which can act synergistically to promote oncogenesis. Our results suggest that Jade-1 may fulfill its tumor suppressor function by inhibiting both the PI3K-AKT and the beta-catenin pathways, which may be important therapeutic targets in renal cancer. |
