title: Wnt5a and Wnt11 inhibit the canonical Wnt pathway and promote cardiac progenitor development via the Caspase-dependent degradation of AKT.
abstract: Wnt proteins regulate cell behavior via a canonical signaling pathway that induces β-catenin dependent transcription. It is now appreciated that Wnt/β-catenin signaling promotes the expansion of the second heart field (SHF) progenitor cells that ultimately give-rise to the majority of cardiomyocytes. However, activating β-catenin can also cause the loss of SHF progenitors, highlighting the necessity of precise control over β-catenin signaling during heart development. We recently reported that two non-canonical Wnt ligands, Wnt5a and Wnt11, act cooperatively to attenuate canonical Wnt signaling that would otherwise disrupt the SHF. While these data reveal the essential role of this anti-canonical Wnt5a/Wnt11 signaling in SHF development, the mechanisms by which these ligands inhibit the canonical Wnt pathway are unclear. Wnt11 was previously shown to inhibit β-catenin and promote cardiomyocyte maturation by activating a novel apoptosis-independent function of Caspases. Consistent with these data, we now show that Wnt5a and Wnt11 are capable of inducing Caspase activity in differentiating embryonic stem (ES) cells and that hearts from Wnt5a(-/-); Wnt11(-/-) embryos have diminished Caspase 3 (Casp3) activity. Furthermore, SHF markers are reduced in Casp3 mutant ES cells while the treatment of wild type ES cells with Caspase inhibitors blocked the ability of Wnt5a and Wnt11 to promote SHF gene expression. This finding was in agreement with our in vivo studies in which injecting pregnant mice with Caspase inhibitors reduced SHF marker expression in their gestating embryos. Caspase inhibition also blocked other Wnt5a/Wnt11 induced effects, including the suppression of β-catenin protein expression and activity. Interestingly, Wnt5a/Wnt11 treatment of differentiating ES cells reduced both phosphorylated and total Akt through a Caspase-dependent mechanism and phosphorylated Akt levels were increased in the hearts Caspase inhibitor treated. Surprisingly, inhibition of either Akt or PI3K in ES cells was an equally effective means of increasing SHF markers compared to treatment with Wnt5a/Wnt11. Moreover, Akt inhibition restored SHF gene expression in Casp3 mutant ES cells. Taken together, these findings suggest that Wnt5a/Wnt11 inhibit β-catenin to promote SHF development through Caspase-dependent Akt degradation.
authors.affiliation: Division of Endocrinology and Metabolism, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA; Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA.
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Bisson
authors.affiliation: Division of Endocrinology and Metabolism, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA; Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA.
authors.forename: Bradley
authors.initials: B
authors.lastname: Mills
authors.affiliation: Division of Endocrinology and Metabolism, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA; Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA.
authors.forename: Jay-Christian
authors.initials: JC
authors.lastname: Paul Helt
authors.affiliation: Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Department of Developmental and Regenerative Medicine, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Zwaka
authors.affiliation: Division of Endocrinology and Metabolism, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA; Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA. Electronic address: Ethan_Cohen@URMC.Rochester.edu.
authors.forename: Ethan David
authors.initials: ED
authors.lastname: Cohen
mesh_terms: D000818:Animals; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006321:Heart / Q000196:embryology*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D009206:Myocardium / Q000166:cytology; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011270:Pregnancy, Animal; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology*; D051153:Wnt Proteins / Q000378:metabolism*; D060449:Wnt Signaling Pathway / Q000502:physiology*; D000071818:Wnt-5a Protein
chemical_list: D051153:Wnt Proteins; D000071818:Wnt-5a Protein; C508957:Wnt11 protein, mouse; C438615:Wnt5a protein, mouse; C514862:Akt1 protein, mouse; D051057:Proto-Oncogene Proteins c-akt; D020169:Caspases
keywords: Akt; Cardiac progenitor cell; Caspase; Heart; Wnt signaling; β-Catenin