SPICULOGENESIS IN THE SEA-URCHIN EMBRYO - STUDIES ON THE SM30 SPICULEMATRIX PROTEIN

When proteins isolated from spicules of Strongylocentrotus purpuratus embryos were examined by western blot analysis, a major protein of app roximately 43 kDa was observed to react with the monoclonal antibody, mAb 1223. Previous studies have established that this antibody recogni zes an asparagine-linked, anionic carbohydrate epitope on the cell sur face glycoprotein, msp130. This protein has been shown to be specifica lly associated with the primary mesenchyme cells involved in assembly of the spicule. Moreover, several lines of evidence have implicated th e carbohydrate epitope in Ca2+ deposition into the growing spicule. Th e 43 kDa, spicule matrix protein detected with mAb 1223 also reacted w ith a polyclonal antibody to a known spicule matrix protein, SM30. Fur ther characterization experiments, including deglycosylation using PNG aseF, two-dimensional electrophoresis, and immunoprecipitation, verifi ed that the 43 kDa spicule matrix protein had a pi of approximately 4. 0, contained the carbohydrate epitope recognized by monoclonal antibod y mAb 1223 and reacted with anti-SM30. Electron microscopy confirmed t he presence of proteins within the demineralized spicule that reacted with mAb 1223 and anti-SM3O. We conclude that the spicule matrix prote in, SM30, is a glycoprotein containing carbohydrate chains similar or identical to those on the primary mesenchyme cell membrane glycoprotei n, msp130.