MicroRNAs regulate sea urchin gut specification and development by targeting components of the Wnt signaling pathway

Nigam, Priya A.
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University of Delaware
microRNAs (miRNAs) are established regulators of cellular signaling and biological processes such as differentiation, apoptosis and proliferation. Previously, our laboratory has demonstrated that miRNAs may regulate endodermal and mesodermal specification (Song et al. , 2012). The canonical Wnt signaling pathway plays a highly conserved role in endodermal specification (Logan et al. , 1999; Wikramanayake et al. , 1998) and proper gut formation (Theodosiou and Tabin, 2003). Therefore, this study tested the hypothesis that miRNAs regulate endodermal specification and gut development in part by regulating conserved components of the canonical Wnt signaling pathway. Using luciferase reporter assays and site-directed mutagenesis of reporter constructs, we demonstrate that miRNAs directly regulate both β-catenin and Dishevelled ( Dsh/Dvl ) post-transcriptionally. Blocking miRNA-mediated regulation of β-catenin by target protector morpholino antisense oligonucleotide (miRNA TP) resulted in: (1) increased β-catenin protein levels, (2) upregulation of β-catenin responsive genes that are involved in endodermal specification, and (3) a structurally and functionally defective gut tissue. In addition, we also observed that inhibition of miRNA-mediated regulation of Dsh/Dvl , an upstream regulator of β-catenin, by use of miRNA TP resulted in an even more severe phenotype of a morphologically thinner gut than treatment with the β-catenin miRNAT TP. In summary, we identified two key components of Wnt signaling to be directly regulated by miRNAs. Importantly, we demonstrated that removal of miRNA-mediated regulation of either β-catenin or Dsh/Dvl in the developing embryo led to defects in endodermally derived gut tissue.