Embryonic nuclear proteins from the sea urchin Strongylocentrotus purpuratus bind in vitro to a cis-acting element that lies upstream of the actin gene CyIIIb and consists of two direct repeats homologous to steroid hormone response elements. This sea urchin element is specifically recognized by transcription factor COUP-TF from HeLa cell nuclear extracts as well. A sea urchin gene homologous to the human COUP-TF1 gene was detected by blot hybridization to S. purpuratus genomic DNA. Screening of a genomic DNA library with a human COUP-TF1 cDNA probe produced overlapping genomic clones that carry the S. purpuratus gene. Our results indicate that the sea urchin homologue has a structure similar to the human COUP-TF1 gene, i.e., conserved intron positions characteristic of this subgroup of steroid hormone receptors. Sequencing of the exons that encode the DNA- and ligand-binding domains of the sea urchin protein revealed 96% and 92% amino acid identity to the domains of the human protein, respectively. Transcripts derived from the S. purpuratus COUP-TF homologous gene were detected in ovarian and embryonic RNAs from various stages.