P3.83 Friday, Jan. 6 Investigating the molecular determinants for polarity in the sea urchin egg WANG, Lingyu*; OOKA, Shioh; POUSTKA, Albert ; WIKRAMANAYAKE, Athula; University of Miami; University of Miami; Max-Planck Institut für Molekulare Genetik, Germany; University of Miami firstname.lastname@example.org
The Dishevelled (Dsh) protein in the Wnt signaling pathway is required for endomesoderm specification in sea urchin embryos. Dsh is highly enriched in a vegetal cortical domain of the unfertilized egg, and during early cleavage stages this protein is selectively activated in vegetal pole blastomeres where it initiates Wnt/beta-catenin signaling. The mechanisms that tether Dsh to the vegetal cortex, and activate this protein in the Wnt pathway during early development are unknown, but this information is critical for understanding early pattern formation in the sea urchin embryo. To identify candidate molecules with potential roles in regulating Dsh function in the early embryo, we carried out two separate molecular screens. To identify all RNAs enriched in the egg cortex, we did a large-scale RNA-seq screen. The mRNAs of most Wnt signaling pathway core components were enriched in the egg cortex. We are currently using in situ hybridization to identify those RNAs that are asymmetrically distributed along the animal vegetal axis. To begin to characterize the Dsh-binding scaffold in the vegetal pole and to identify Dsh-interacting proteins in 16-cell stage micromeres, we carried out co-immunoprecipitation (Co-IP) experiments. These Co-IP studies have identified several proteins that potentially interact with Dsh in the cortex and in 16-cell stage embryos. We will describe the functional roles of these proteins in tethering Dsh to the vegetal cortex, and in "activating" Dsh in the Wnt/beta-catenin pathway.