Dorsal activity of maternal squint is mediated by a non-coding function of the RNA

• Main Author: Lim, Shi Min
• Other Authors: School of Biological Sciences
• Format: Theses and Dissertations
• Language: English
• Published: 2014
• Subjects: DRNTU::Science::Biological sciences::Genetics
• Online Access: https://hdl.handle.net/10356/58582
• Institution: Nanyang Technological University

The earliest steps of axis formation in zebrafish are thought to be regulated by maternal factors, for instance, that activate Wnt/β-Catenin signaling to specify the dorsal axis. It was previously shown that asymmetric localization of maternal transcripts of the conserved zebrafish TGF-β factor, Squint (Sqt/Nodal-related 1, Ndr1), in 4-cell stage embryos predicts the position of the future embryonic dorsal, preceding dorsal nuclear accumulation of β-Catenin (Gore et al., 2005). The nodal genes are classically involved in mesoderm and endoderm formation, left-right axial patterning, neural patterning and stem cell maintenance. However, cell ablations and antisense oligonucleotides that deplete Sqt lead to dorsal deficiencies, suggesting that localized maternal sqt RNA functions in dorsal specification. Due to discrepancies between the genetic nodal mutants and the ablations/antisense knock-down results, the function and mechanism of maternal sqt was debated (Bennett et al., 2007; Gore et al., 2007). In this study, I show that sqt RNA has activity independent of Sqt protein in dorsal specification. Surprisingly, over-expression of mutant/non-coding sqt RNA and particularly, the sqt 3’UTR, leads to increased number of β-Catenin-positive nuclei and expands dorsal gene expression. Dorsal activity of sqt RNA requires Wnt/β-Catenin but not co-receptor Oep-dependent Nodal signaling, explaining the discrepancy between the nodal signaling mutants and the morpholino phenotypes. Also, depletion of maternal sqt RNA abolishes nuclear β-Catenin, providing the mechanism for the loss of dorsal in the morphants, and places activity of maternal sqt RNA upstream of β-Catenin. Remarkably, this loss of early dorsal gene expression can be rescued by the sqt 3’UTR sequences. My findings identify new non-coding functions for the nodal genes, and support a model wherein sqt RNA acts as a scaffold to bind and deliver/sequester maternal factors to future embryonic dorsal.