Sea urchin early histone genes are active in preblastula embryos; late histone genes are maximally expressed during subsequent stages of embryogenesis. We used the Xenopus laevis oocyte to assay for trans-acting factors involved in this differential regulation. Sea urchin nuclear proteins were prepared by extracting gastrula-stage chromatin successively with 0.45, 1, and 2 M NaCl. We injected three fractions into oocytes along with plasmids bearing sea urchin early and late H2b histone genes. While neither the 0 to 0.45 M nor the 1 to 2 M salt fraction affected H2b gene expression, the 0.45 to 1 M salt fraction stimulated early and late H2b mRNA levels significantly. Late H2b gene expression was stimulated preferentially when the early and late genes were coinjected into the same oocytes. This extract did not stimulate the accumulation of transcripts of injected herpesvirus thymidine kinase genes or of the sea urchin Spec 1 gene, suggesting that the stimulatory activity is not a general transcription factor. We localized the DNA sequence required for the stimulatory effect to a region of the late H2b gene located between -43 and +62 relative to the transcription start site. A component of the 0.45 to 1 M salt wash fraction specifically bound to the 105-base-pair late gene DNA sequence and to the corresponding early gene fragment. The abundance of this binding activity decreased on a per genome basis during early development of the sea urchin.