22 décembre 19351935/12/22 (N503).Appartient à l’ensemble documentaire : PACA

Anita Khadka (4787565)

Marina Martínez-Bartolomé (4787559)

Ryan Range (234426)

Stephanie Burr (4787562)

EvoDevo

Gallica, Bibliotheque Numerique

Les Tablettes d'Avignon et de Provence : revue hebdomadaire illustrée de propagande régionaliste par Avignon, centre de tourisme

Abstract The anterior neuroectoderm (ANE) in many deuterostome embryos (echinoderms, hemichordates, urochordates, cephalochordates, and vertebrates) is progressively restricted along the anterior–posterior axis to a domain around the anterior pole. In the sea urchin embryo, three integrated Wnt signaling branches (Wnt/β-catenin, Wnt/JNK, and Wnt/PKC) govern this progressive restriction process, which begins around the 32- to 60-cell stage and terminates by the early gastrula stage. We previously have established that several secreted Wnt modulators of the Dickkopf and secreted Frizzled-related protein families (Dkk1, Dkk3, and sFRP-1/5) are expressed within the ANE and play important roles in modulating the Wnt signaling network during this process. In this study, we use morpholino and dominant-negative interference approaches to characterize the function of a novel Frizzled-related protein, secreted Frizzled-related protein 1 (sFRP-1), during ANE restriction. sFRP-1 appears to be related to a secreted Wnt modulator, sFRP3/4, that is essential to block Wnt signaling and establish the ANE in vertebrates. Here, we show that the sea urchin sFRP3/4 orthologue is not expressed during ANE restriction in the sea urchin embryo. Instead, our results indicate that ubiquitously expressed maternal sFRP-1 and Fzl1/2/7 signaling act together as early as the 32- to 60-cell stage to antagonize the ANE restriction mechanism mediated by Wnt/β-catenin and Wnt/JNK signaling. Then, starting from the blastula stage, Fzl5/8 signaling activates zygotic sFRP-1 within the ANE territory, where it works with the secreted Wnt antagonist Dkk1 (also activated by Fzl5/8 signaling) to antagonize Wnt1/Wnt8–Fzl5/8–JNK signaling in a negative feedback mechanism that defines the outer ANE territory boundary. Together, these data indicate that maternal and zygotic sFRP-1 protects the ANE territory by antagonizing the Wnt1/Wnt8–Fzl5/8–JNK signaling pathway throughout ANE restriction, providing precise spatiotemporal control of the mechanism responsible for the establishment of the ANE territory around the anterior pole of the sea urchin embryo

Anita Khadka

Marina Martínez-Bartolomé

Stephanie D. Burr

Ryan C. Range

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A novel gene’s role in an ancient mechanism: secreted Frizzled-related protein 1 is a critical component in the anterior–posterior Wnt signaling network that governs the establishment of the anterior neuroectoderm in sea urchin embryos

Additional file 2: Figure S2. sFRP-1 morpholino control experiments. (A) The expression of ANE regulatory factors six3, foxq2, nkx3.2, and sfrp-1/5 are severely downregulated in embryos injected with sFRP-1 morpholino 2 designed to interfere with the splicing at the first exon–intron boundary. (B) Low levels of sFRP-1 expression (1 μg/μL) rescue embryos injected with sFRP-1. MO1. The number of embryos examined that show the representative phenotypes depicted is indicated in each panel. sFRP-1 MO1 does not bind to exogenous sfrp-1 mRNA. (C) Diagram of the intron–exon organization of sfrp-1 pre-mRNA. Primers used to characterize the mRNA products (orange arrows). Position of the target sequence for the morpholino (red bar). MO, morpholino. (D) Efficacy control for the sFRP-1 splice-blocking morpholino. PCR analysis of control glycerol injected and embryos injected with a sFRP-1 splice-blocking morpholino (sFRP-1 MO2 in methods). Expected control PCR product size for sfrp-1 = 100 bps; no PCR product expected from sFRP-1 MO2-injected embryos

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Additional file 1: Figure S1. Phylogenetic analysis of sea urchin Frizzled-like cysteine-rich domains and sfrp3/4 expression during ANE restriction. (A) Fzl-like cysteine-rich domain maximum likelihood analysis. The Bayesian tree was built using only Fzl-CRD amino acid sequences. Each Fzl-CRD orthologue is highlighted by a different color. Those proteins that contain more than one Fzl-CRD are labeled with a numbered CRD corresponding to its position in the protein. (B) qPCR measurements from three different cultures of embryos showing the approximate number of sfrp3/4 transcripts per embryo at 0 hpf, 60-cell, 120-cell, hatched blastula, and mesenchyme blastula stages. The y axis shows the approximate number of transcripts per embryo based on the Ct value of z12 transcripts. The absolute concentrations of z12 transcripts are known at each stage [60]. FE, fertilized egg; EB, early blastula; B, blastula; MB, mesenchyme blastula

MOESM1 of A novel gene’s role in an ancient mechanism: secreted Frizzled-related protein 1 is a critical component in the anterior–posterior Wnt signaling network that governs the establishment of the anterior neuroectoderm in sea urchin embryos

Additional file 3: Figure S3. Spatiotemporal expression analysis of Dkk1, Fzl5/8, and sFRP-1. (A) Whole-mount in situ hybridization analysis of dkk1 expression during ANE restriction. (B) sfrp-1 (a) and fzl5/8 (b) mRNA transcripts are expressed in remarkably similar territories mesenchyme blastula stage embryos (24 hpf)

MOESM3 of A novel gene’s role in an ancient mechanism: secreted Frizzled-related protein 1 is a critical component in the anterior–posterior Wnt signaling network that governs the establishment of the anterior neuroectoderm in sea urchin embryos

https://figshare.com/articles/MOESM3_of_A_novel_gene_s_role_in_an_ancient_mechanism_secreted_Frizzled-related_protein_1_is_a_critical_component_in_the_anterior_posterior_Wnt_signaling_network_that_governs_the_establishment_of_the_anterior_neuroectoderm_in_sea_urchin_embryos/5812488

Les Tablettes d'Avignon et de Provence : revue hebdomadaire illustrée de propagande régionaliste par Avignon, centre de tourisme

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