title: A phenotypic screen in zebrafish identifies a novel small-molecule inducer of ectopic tail formation suggestive of alterations in non-canonical Wnt/PCP signaling.
abstract: Zebrafish have recently emerged as an attractive model for the in vivo bioassay-guided isolation and characterization of pharmacologically active small molecules of natural origin. We carried out a zebrafish-based phenotypic screen of over 3000 plant-derived secondary metabolite extracts with the goal of identifying novel small-molecule modulators of the BMP and Wnt signaling pathways. One of the bioactive plant extracts identified in this screen - Jasminum gilgianum, an Oleaceae species native to Papua New Guinea - induced ectopic tails during zebrafish embryonic development. As ectopic tail formation occurs when BMP or non-canonical Wnt signaling is inhibited during the tail protrusion process, we suspected a constituent of this extract to act as a modulator of these pathways. A bioassay-guided isolation was carried out on the basis of this zebrafish phenotype, identifying para-coumaric acid methyl ester (pCAME) as the active compound. We then performed an in-depth phenotypic analysis of pCAME-treated zebrafish embryos, including a tissue-specific marker analysis of the secondary tails. We found pCAME to synergize with the BMP-inhibitors dorsomorphin and LDN-193189 in inducing ectopic tails, and causing convergence-extension defects in compound-treated embryos. These results indicate that pCAME may interfere with non-canonical Wnt signaling. Inhibition of Jnk, a downstream target of Wnt/PCP signaling (via morpholino antisense knockdown and pharmacological inhibition with the kinase inhibitor SP600125) phenocopied pCAME-treated embryos. However, immunoblotting experiments revealed pCAME to not directly inhibit Jnk-mediated phosphorylation of c-Jun, suggesting additional targets of SP600125, and/or other pathways, as possibly being involved in the ectopic tail formation activity of pCAME. Further investigation of pCAME's mechanism of action will help determine this compound's pharmacological utility.
authors.affiliation: Laboratory for Molecular Biodiscovery, Department of Pharmaceutical and Pharmacological Sciences, University of Leuven, Leuven, Belgium.
authors.forename: Evelien
authors.initials: E
authors.lastname: Gebruers
authors.affiliation: Laboratory for Molecular Biodiscovery, Department of Pharmaceutical and Pharmacological Sciences, University of Leuven, Leuven, Belgium ; Faculty of Chemistry Sciences, School of Biochemistry and Pharmacy, University of Cuenca, Cuenca, Ecuador ; Chemical Biology Group, Luxembourg Centre for Systems Biomedicine, University of Luxembourg, Esch-sur-Alzette, Luxembourg.
authors.forename: María Lorena
authors.initials: ML
authors.lastname: Cordero-Maldonado
authors.affiliation: Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, Scotland.
authors.forename: Alexander I
authors.initials: AI
authors.lastname: Gray
authors.affiliation: Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, Scotland.
authors.forename: Carol
authors.initials: C
authors.lastname: Clements
authors.affiliation: Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, Scotland.
authors.forename: Alan L
authors.initials: AL
authors.lastname: Harvey
authors.affiliation: Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, Scotland.
authors.forename: Ruangelie
authors.initials: R
authors.lastname: Edrada-Ebel
authors.affiliation: Laboratory for Molecular Biodiscovery, Department of Pharmaceutical and Pharmacological Sciences, University of Leuven, Leuven, Belgium.
authors.forename: Peter A M
authors.initials: PA
authors.lastname: de Witte
authors.affiliation: Laboratory for Molecular Biodiscovery, Department of Pharmaceutical and Pharmacological Sciences, University of Leuven, Leuven, Belgium ; Chemical Biology Group, Luxembourg Centre for Systems Biomedicine, University of Luxembourg, Esch-sur-Alzette, Luxembourg.
authors.forename: Alexander D
authors.initials: AD
authors.lastname: Crawford
authors.affiliation: Laboratory for Molecular Biodiscovery, Department of Pharmaceutical and Pharmacological Sciences, University of Leuven, Leuven, Belgium.
authors.forename: Camila V
authors.initials: CV
authors.lastname: Esguerra
mesh_terms: D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D003373:Coumaric Acids / Q000737:chemistry / Q000494:pharmacology*; D004353:Drug Evaluation, Preclinical; D004625:Embryo, Nonmammalian / Q000196:embryology*; D031660:Jasminum / Q000737:chemistry*; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D013623:Tail / Q000196:embryology*; D060449:Wnt Signaling Pathway / Q000187:drug effects*; D015027:Zebrafish / Q000196:embryology*; D029961:Zebrafish Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D019485:Bone Morphogenetic Proteins; D003373:Coumaric Acids; C554430:LDN 193189; D011720:Pyrazoles; D011743:Pyrimidines; D029961:Zebrafish Proteins; C516138:dorsomorphin; C057918:4-coumaric acid methyl ester