MOLECULAR-CLONING OF A CDNA THAT ENCODES THE PRECURSOR TO SEVERAL EXOGASTRULA-INDUCING PEPTIDES, EPIDERMAL-GROWTH-FACTOR-RELATED POLYPEPTIDES OF THE SEA-URCHIN ANTHOCIDARIS-CRASSISPINA

Complementary DNA clones for exogastrula-inducing peptides (EGIPs) of the sea urchin Anthocidaris crassispina, which are related to epiderma l growth factor (EGF), were obtained from a cDNA library of late gastr ula embryos using, as probe, the partial cDNA for one of the EGIP (EGI P-D) obtained by the reverse-transcription PCR method. The longest cDN A was composed of 1662 bp, and encoded a protein of approximately 36 k Da with a region that resembled a signal sequence. The deduced protein contains the sequences of EGIP-C, EGIP-D, and EGIP-A in that order, f ollowed by the sequence for an unidentified EGIP-like polypeptide. Whe n expressed in Escherichia coli as a fusion protein with beta-galactos idase, the product for the cDNA was specifically recognized by a rabbi t antibody raised against EGIP-D that had been purified from embryos. Characteristic amino acid residues were found around the N-terminus an d the C-terminus of each EGIP sequence, suggesting a specific processi ng mechanism for the generation of the individual EGIPs from the precu rsor. RNA-blot analysis revealed the presence of EGIP mRNA in unfertil ized eggs. The level of this mRNA decreased gradually after fertilizat ion, began to increase dramatically after the onset of gastrulation, a nd continued to increase through the pluteus stage. Genomic Southern-b lot analysis suggested that this gene is present as a single copy. A h omology search showed that the EGIP cDNA has a similarity to the cDNA for SpEGF2 which was cloned as a gastrula-specific gene in another sea urchin, Strongylocentrotus purpuratus.