Expression of vitellogenin, the yolk protein precursor, is strictly regulated during development. In previous studies on a variety of organisms, vitellogenin gene expression has been shown to be restricted to one or two tissues in adult female animals. In this report we show that, in contrast, sea urchin vitellogenin is synthesized in both females and males. To identify sea urchin vitellogenin, we raised antibodies specific for the major yolk protein. We show here that a 155-kDa polypeptide, immunoprecipitable by the antibody to the major yolk protein, is synthesized in the intestines of female and male sea urchins and also in ovaries and testes. This 155-kDa polypeptide is converted to a 195-kDa vitellogenin in each of these tissues; further modification to yield the 180-kDa major yolk protein occurs only in the ovary. We have also identified a vitellogenin cDNA clone and used it to study vitellogenin mRNA production. An abundant 5.1-kilobase mRNA was found in the tissues containing vitellogenin. Our results suggest that vitellogenin may serve the following two functions in sea urchins: its classical role as a yolk protein precursor and an unidentified function required by adults of both sexes.