Essentials Polyphosphate (polyP) activates mTOR but its role in Wnt/ß-catenin signaling is not known. PolyP-mediated cyclin D1 expression (ß-catenin target gene) was monitored in endothelial cells. PolyP and boiled platelet-releasates induced the expression of cyclin D1 by similar mechanisms. PolyP establishes crosstalk between mTOR and Wnt/ß-catenin signaling in endothelial cells. SUMMARY: Background Inorganic polyphosphate (polyP) elicits intracellular signaling responses in endothelial cells through activation of mTOR complexes 1 and 2. Glycogen synthase kinase 3 (GSK-3) is known to be a negative regulator of mTOR and Wnt/ß-catenin signaling pathways. Objective The objective of this study was to investigate the effect of polyP on the expression, degradation and subcellular localization of the Wnt/ß-catenin target gene, cyclin D1, in endothelial cells. Methods Regulation of cyclin D1 expression, phosphorylation and subcellular localization by polyP or platelet releasates was monitored in the absence and presence of pharmacological inhibitors and/or siRNA for specific molecules of the upstream mTOR/Wnt/ß-catenin signaling network by established methods. Results Both synthetic polyP and boiled-platelet releasates induced the phosphorylation-dependent inactivation of GSK-3, thereby increasing the expression and nuclear localization, but inhibiting the degradation of cyclin D1. Inhibitors of mTORC1 (PI3K, AKT, PLC, PKC), rapamycin and siRNA for raptor (mTORC1-specific component) and ß-catenin, all inhibited polyP-mediated regulation of cyclin D1 expression, phosphorylation and subcellular localization in endothelial cells. The signaling effect of polyP was effectively inhibited by the recombinant extracellular domain of the receptor for advanced glycation end products (RAGE) and/or by the RAGE siRNA. Specific pharmacological inhibitors and siRNA knockdown of ERK1/2 and NF-κB pathways indicated that polyP-mediated cyclin D1 expression and nuclear localization are IKKɑ and ERK1/2 dependent, whereas its inhibitory effect on phosphorylation-dependent degradation of cyclin D1 is IKKß-dependent. Conclusion We conclude that a RAGE-dependent polyP-mediated crosstalk between mTOR and the GSK-3/Wnt/ß-catenin signaling network can modulate important physiological processes in endothelial cells.