Southern blot analysis of DNA derived from sea urchin, Lytechinus variegatus, that had been cleaved by EcoRI or HindIII revealed a major strong hybridization band using a v-ets probe. The band obtained was constructed from a charon 28 library from DNA fragments of this size. A phage (12E3) containing sequences hybridizing to the E26 v-ets probe was isolated from this library and the ets-homologous region was sequenced by the dideoxynucleotide chain-termination method. A highly homologous sequence to E26 v-ets was found; this region is the same one that also corresponds to the human (Hu-ets-2) homologous sequences defined in our lab. The sea urchin homology with v-ets begins at a consensus splice acceptor sequence and ends at the point where it is known that v-ets and Hu-ets homology diverge. Ninety-one out of 97 (or 94%) predicted amino acids share identity between the sea urchin c-ets and E26 v-ets over their region of homology. A somewhat weaker homology with the Hu-ets-2 sequences continues beyond this point for 13 more codons, ending at a common termination codon. Methods for culturing the embryos of sea urchin and Xenopus larvis have been established. A single 6.8 kb ets-related RNA was observed by Northern blot analysis from the unfertilized egg stage until the blastula stage of development in the sea urchin embryos. The maximal level of expression occurred in the early stages of embryonic sea urchin development (16 cells to morula stage). Western blot and immunoprecipitation analysis of sea urchin embryo protein extracts revealed a 72 kDa band that is identifiable by anti-human ets-2 peptide antibody. Microinjection of antibody (anti-ets-2) into sea urchin embryos has been started in order to find some clues to the ets gene functions in these cells. Several positive clones have already been found from screening of the Xenopus larvis DNA library with the v-ets probe. Both cDNA library constructions have been started.
