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Rab11a-recycling endosomes function to regulate cell signaling during retinal neurogenesis in zebrafish

ABSTRACT: We report the transcriptional changes associated with inhibition of recycling endosome function within the developing zebrafish retina. Recycling endosome function was inhibited through transgenic expression of a Rab11a dominant negative protein within the developing retinal progenitor cells using UAS::mCherry-Rab11aS25N; vsx2::Gal4 transgenic zebrafish. Changes in gene expression, compared to vsx2:Gal4 controls were profiled through bulk RNA-sequencing experiments of dissected zebrafish retinas. Gene expression changes resulting from inhibition of recycling endosome function were then compared to experiments in which canonical signaling pathways were modulated, including activation of Notch-signaling (over-expression of the constitutively active Notch1a intracellular domain; UAS::myc-NICD1a; vsx2::Gal4), activation of Wnt-signaling (over-expression of Wnt2b; UAS::EGFP-Wnt2b;; vsx2::Gal4), activation of Hippo-signaling (over-expression of nuclear retained YapS87A; dRED::UAS::YapS87A; vsx2::Gal4), and inhibition of mTor signaling (through treatment of zebrafish embryos with Torrin).

ORGANISM(S): Danio rerio

PROVIDER: GSE154895 | GEO | 2021/02/22

REPOSITORIES: GEO

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Project description:Distinct shaping of the upper versus lower facial skeleton is essential for function of the vertebrate jaw and middle ear, yet the cellular mechanisms by which this occurs have remained unclear. Here, we show that Endothelin1 (Edn1) signaling accelerates mesenchymal condensation and subsequent cartilage formation in the lower face through antagonism of Jagged-Notch signaling and Prrx1 transcription factors. A genomic analysis of facial skeletal precursors in mutants and overexpression embryos reveals that Jagged-Notch signaling represses genes that are strongly induced as pharyngeal arch neural crest-derived cells begin skeletal differentiation. In wild types, initial Jagged-Notch repression dorsally ensures that barx1+ condensations and cartilage differentiation occur first in ventral-intermediate zones of the pharyngeal arches. Reduced Jagged-Notch signaling results in an expansion of pre-cartilage condensations in the upper face, with loss of barx1 partially restoring dorsal cartilage shapes in jag1b mutants. Further, by studying new mutants for zebrafish prrx1a and prrx1b, we find that Prrx1 genes function in parallel to Jagged-Notch signaling to restrict the formation of dorsal barx1+ pre-cartilage condensations. Consistently, combined losses of jag1b and prrx1a/b robustly rescue ventral barx1+ condensations and lower facial cartilage development in edn1 mutants. Together, our work suggests that Edn1 works through parallel inhibition of Jagged-Notch and Prrx1 pathways to promote an earlier and more extensive establishment of cartilage condensations in the lower face. We performed RNAseq on FACS-sorted neural crest-derived pharyngeal arch cells (fli1a:GFP; sox10:DsRed double positive) from wild-type embryos at 3 different stages (20, 28, and 36 hours post fertilization) and embryos with altered levels of Edn1 and Notch signaling (edn1 mutants and hsp70I:Gal4; UAS:Edn1 transgenics; jag1b mutants, dibenzazepine-treated embryos, and hsp70I:Gal4; UAS:NICD transgenics. We also sequenced RNA from heat-shocked UAS:Edn1+ and hsp70I:Gal4+ transgenics and jag1b+/+ controls.

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Rab11a-recycling endosomes function to regulate cell signaling during retinal neurogenesis in zebrafish

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