Histological distribution of FR-1, a cyclic RGDS-peptide, binding sites during early embryogenesis, and isolation and initial characterization of FR-1 receptor in the sand dollar embryo.
A fibronectin-related synthetic cyclic H-Cys-Arg-Gly-Asp-Ser-Pro-Ala-Ser-Ser-Cys-OH (RGDSPASS) peptide (FR-1) binding site in the embryo of the sand dollar Clypeaster japonicus was specified using dansyl-labeled FR-1 (Dns-FR-1) and horseradish peroxidase-labeled FR-1, and an FR-1 receptor was isolated using FR-1-affinity column chromatography. The FR-1 introduced to the blastocoel of blastulae inhibited primary mesenchyme cell (PMC) migration in mesenchyme blastulae, and complete gastrulation and spicule differentiation in gastrulae. The Dns-FR-1 bound to the entire basal side of the ectoderm in mesenchyme blastulae, and then restricted to the basal side of the ectoderm at the apical tuft region and the vegetal hemisphere in early gastrulae. The cytoplasm of the archenteron also bound to Dns-FR-1. In PMC, Dns-FR-1 bound to the nucleus and cytoplasmic reticular features. In unfertilized eggs, Dns-FR-1 bound to the entire cytoplasm, particularly to the oval-shaped granules and the nuclear envelope, but only to the cytoplasm after fertilization. Relative molecular mass (Mr) of the FR-1-binding protein was 240 kDa under non-reducing conditions and 57 kDa under reducing conditions. The FR-1 receptor protein bound anti-sea urchin integrin (Spl) betaL subunit antibodies raised against the embryos of Strongylocentrotus purpuratus. Immunohistochemistry showed that the antibody binding site was similar to the histochemical distribution of Dns-FR-1. However, Mr of the FR-1 receptor is distinctively larger than that of the Spl betaL subunit.
Katow
H
H
Biology Laboratory, College of Science, Rikkyo University, Tokyo, Japan.
Yamamoto
Y
Y
Sofuku
S
S
D000818:Animals; D001665:Binding Sites; D002846:Chromatography, Affinity; D003208:Concanavalin A / Q000378:metabolism; D003619:Dansyl Compounds; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D007163:Immunosorbent Techniques; D066298:In Vitro Techniques; D008958:Models, Molecular; D009024:Morphogenesis; D009842:Oligopeptides / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D010063:Ovum / Q000378:metabolism; D012617:Sea Urchins / Q000196:embryology*
D003619:Dansyl Compounds; D005456:Fluorescent Dyes; D009842:Oligopeptides; C106780:cyclic(arginyl-glycyl-aspartyl-seryl-prolyl-alanyl-seryl-serine); D003208:Concanavalin A; C046617:arginyl-glycyl-aspartyl-serine