A 70 kD microtubule‐binding protein from starfish eggs: Purification, characterization, and localization during meiosis and mitosis

AbstractA microtubule‐binding protein was purified from eggs of the starfish, Asterias amurensis, through several steps of purification including the taxol‐dependent procedure [Vallee, 1982, J. Cell Biol. 92:435–442]. This protein consists of a single polypeptide chain having an apparent molecular mass of 70 kD determined by SDS‐PAGE. The 70 kD protein was identified as a unique microtubulebinding protein, judging from electrophoretic mobility, cleavage pattern by limited proteolysis, heat stability, and immunocrossreactivity. The 70 kD protein binds to brain and egg microtubules. It does not promote assembly of brain tubulin, but promotes that of egg tubulin in vitro in a concentration‐dependent manner.Using indirect immunofluorescence and immunoelectron microscopy with the anti‐70 kD protein antibody, we analyzed the cellular localization of the 70 kD protein in starfish oocytes and eggs during both meiotic maturation (meicsis) and first cleavage (mitosis). Immunofluorescence studies showed that the 70 kD protein localized on microtubule structures spread widely throughout the cytoplasm, the sperm aster, and the microtubules making up the mitotic apparatus through both meiosis and mitosis. The antibody, however, did not recognize sperm axonemes. These results were confirmed by immunoelectron microscopy. Using a colloidal gold technique, the 70 kD protein was localized along the microtubules in vivo.This 70 kD protein is the first microtubule‐binding protein that has been shown to localize along the microtubules in oocytes and eggs throughout meiosis and mitosis and to promote microtubule assembly. The 70 kD protein may be involved in the dynamic changes of microtubule structures occurring within oocytes and eggs.