Apextrin, a Novel Extracellular Protein Associated with Larval Ectoderm Evolution in Heliocidaris erythrogramma

Abstract

During the evolution of direct development in the sea urchin Heliocidaris erythrogramma major modifications occurred, which allowed the precocious formation of adult-specific structures and led to a novel larval body that surrounds these structures. The HeET-1 gene was isolated in a differential screen for transcripts enriched in the early embryos of H. erythrogramma relative to those of its indirect-developing congener, H. tuberculata. HeET-1 was unique among the three genes found in that no homologous transcript was detected in H. tuberculata total embryonic RNA blots. To verify this apparently extreme differential expression of the HeET-1 genes in Heliocidaris, we isolated the HeET-1 homologue from H. tuberculata genomic DNA and used it to probe blots of poly(A)+ RNA prepared from H. tuberculata embryos. It is expressed in H. tuberculata embryos at levels undetectable by this technique. The predicted amino acid sequence of HeET-1 suggested that it encodes a novel secreted protein. To assess the function of HeET-1, we raised polyclonal antisera to the HeET-1-encoded protein. We find that it is present in eggs in a type of secretory vesicle and that this maternal pool is gradually secreted after fertilization. As cells acquire apical–basal polarity in the blastula the protein becomes localized to the apical extracellular matrix, leading us to name the protein apextrin. The apical extracellular localization of apextrin is maintained in the columnar cells of the larval ectoderm until their internalization at metamorphosis. Ingressing mesenchyme cells rapidly endocytose apextrin upon leaving the vegetal plate. Comparison with fibropellin III, an apical lamina component, suggests that apextrin is an extracellular protein that is in tighter association with the plasma membrane than is the hyalin layer or apical lamina. We propose that apextrin is involved in apical cell adhesion and that its high level of expression may represent an adaptive cooption necessary for strengthening the large H. erythrogramma embryo.