TRANSIENT APPEARANCE OF STRONGYLOCENTROTUS-PURPURATUS OTX IN MICROMERE NUCLEI - CYTOPLASMIC RETENTION OF SPOTX POSSIBLY MEDIATED THROUGH ANALPHA-ACTININ INTERACTION

At the 16-cell stage, the sea urchin embryo is partitioned along the a nimal-vegetal axis into eight mesomeres, four macromeres, and four mic romeres. The micromeres, unlike the other blastomeres, are autonomousl y specified to produce skeletogenic mesenchymal cells and are also req uired to induce the vegetal-plate territory. A long-held belief is tha t micromeres inherit localized maternal determinants that endow them w ith their cell autonomous behavior and inducing capabilities. Here, we present evidence that an orthodenticle-related protein, SpOtx appears transiently in nuclei of micromeres but not in nuclei of mesomeres an d macromeres. At later stages of development, SpOtx was translocated i nto nuclei of all cells. To address the possibility that SpOtx was ret ained in the cytoplasm at early developmental stages, we searched for cytoplasmic proteins that interact with SpOtx. A proline-rich region o f SpOtx resembling an SH3-binding domain was used to screen an embryo cDNA expression library, and a cDNA clone was isolated and shown to be alpha-actinin. A yeast two-hybrid analysis showed a specific interact ion between the proline-rich region of SpOtx and a putative SH3 domain of the sea urchin alpha-actinin. Because micromeres lack an actin-bas ed cytoskeleton, the results suggested that, at the vegetal pole of th e 16-cell stage embryo, SpOtx was translocated into micromere nuclei, whereas in other blastomeres SpOtx was actively retained in the cytopl asm by binding to alpha-actinin. The transient appearance of SpOtx in micromere nuclei may be associated with the specification of micromere cell fate. (C) 1996 Wiley-Liss, Inc.