Suppression of cyclic GMP-dependent protein kinase is essential to the Wnt/cGMP/Ca2+ pathway

Novel downstream effectors sensing changes in intracellular concentrations of Ca(2+) and cyclic GMP in response to activation of the Wnt/Frizzled-2 pathway were sought. Activation of Frizzled-2 suppressed protein kinase G activity, while activating NF-AT-dependent transcription. Each of these responses was abolished by pertussis toxin and by knock-down of the expression of either Galphat2 or Galphao. Activation of NF-AT-dependent transcription in response to Wnt5a stimulation was suppressed by activation of protein kinase G and by buffering intracellular Ca(2+). Elevation of intracellular cyclic GMP either by inhibition of cyclic GMP phosphodiesterase or by addition of 8-bromo-cyclic GMP was shown to activate protein kinase G, to block Ca(2+) mobilization as well as to markedly attenuate activation of NF-AT-dependent transcription in response to Wnt5a stimulation. Chemical inhibition of protein kinase G by Rp-8-pCPT-cGMP, conversely, was shown to provoke increased NF-AT gene transcription and Ca(2+) mobilization, in the absence of Wnt stimulation. Protein kinase G is shown to be a critical downstream effecter of the non-canonical Wnt-Frizzled-2/cGMP/Ca(2+) pathway.