title: PTH/cAMP/PKA signaling facilitates canonical Wnt signaling via inactivation of glycogen synthase kinase-3beta in osteoblastic Saos-2 cells.
abstract: Although the intermittent administration of PTH is known to stimulate the bone formation, the underlying mechanisms are not fully understood. Here we investigated the crosstalk between PTH/cAMP signaling and canonical Wnt signaling using the human osteoblastic cell line Saos-2. Treatment with PTH or forskolin, an activator of adenylate cyclase, facilitated T-cell factor (TCF)-dependent transactivation in a dose-dependent manner, which was abolished by pre-treatment with a PKA inhibitor, H89. Wnt3a and forskolin synergistically increased the TCF-dependent transactivation. Interestingly, intermittent treatment with PTH enhanced the TCF-dependent transactivation more profoundly than continuous treatment. In addition to the effects on TCF-dependent reporter activity, treatment with PTH or forskolin resulted in the increased expression of endogenous targets of Wnts, Wnt-induced secreted protein 2 (WISP2) and naked cuticle 2 (NKD2). We then investigated the convergence point of PTH/cAMP signaling and the canonical Wnt pathway. Western blotting demonstrated that GSK-3beta was rapidly phosphorylated at Ser(9) on treatment with PTH or forskolin, leading to its inactivation. Moreover, overexpression of a constitutively active mutant of GSK-3beta abolished the TCF-dependent transactivation induced by forskolin. On the other hand, overexpression of the Wnt antagonist Dickkopf-1 (DKK1) failed to cancel the effects of forskolin on the canonical Wnt pathway. Interestingly, treatment with Wnt3a markedly reduced the forskolin-induced expression of receptor activator of NF-kappaB ligand (RANKL), a target gene of PTH/cAMP/PKA. These results suggest that cAMP/PKA signaling activates the canonical Wnt pathway through the inactivation of GSK-3beta, whereas Wnt signaling might inhibit bone resorption through a negative impact on RANKL expression in osteoblasts.
authors.affiliation: Department of Bone and Mineral Research, Osaka Medical Center and Research Institute for Maternal and Child Health, Osaka 594-1101, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Suzuki
authors.forename: Keiichi
authors.initials: K
authors.lastname: Ozono
authors.forename: Takuo
authors.initials: T
authors.lastname: Kubota
authors.forename: Hiroki
authors.initials: H
authors.lastname: Kondou
authors.forename: Kanako
authors.initials: K
authors.lastname: Tachikawa
authors.forename: Toshimi
authors.initials: T
authors.lastname: Michigami
mesh_terms: D001862:Bone Resorption; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D010281:Parathyroid Hormone / Q000378:metabolism / Q000494:pharmacology*; D053245:RANK Ligand / Q000096:biosynthesis*; D020239:Receptor Cross-Talk / Q000502:physiology*; D015398:Signal Transduction; D051153:Wnt Proteins / Q000378:metabolism* / Q000494:pharmacology; D060508:Wnt3 Protein; D060509:Wnt3A Protein
chemical_list: D010281:Parathyroid Hormone; D053245:RANK Ligand; C505688:TNFSF11 protein, human; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; D000242:Cyclic AMP; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D017868:Cyclic AMP-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3