Evolution of a novel nuclear receptor subfamily with emphasis on the member from the Pacific oyster Crassostrea gigas

Nuclear receptors (NRs) belong to the transcription factor superfamily that regulates development, homeostasis, differentiation, and reproduction in metazoans via control of gene expression. Recently, rapid advances in genome projects on various metazoans have provided new opportunities for studying the evolution and function of NRs. Typically structured NRs are divided into six subfamilies. Here, the gene for a typically structured NR (CgNR8A1) was cloned from the Pacific oyster Crassostrea gigas. However, this novel receptor could not be assigned to a known NR subfamily. By data mining, nine other CgNR8A1 gene homologs were identified in metazoans such as cnidarians, mollusks, annelids, echinoderms, hemichordates, and cephalochordates. Phylogenetic analysis showed that these receptors belonged to a novel NR subfamily, hereafter designated as NR8. Evolutionary analysis revealed that the NR8 subfamily was phylogenetically the third-oldest NR subfamily, and it originated from a common ancestor of Eumetazoa; several gene loss events occurred independently in ancestors of vertebrates, ecdysozoans, and platyhelminths, which do not have NR8 members. Furthermore, the function of CgNR8A1 was investigated to provide an insight into the functions of this novel NR subfamily. A nuclear localization signal peptide, GKHRNKKPRLD, was identified in CgNR8A1, and a recombinant full-length protein of CgNR8A1 was localized in the nuclei of HeLa cells. The mRNA expression profile of CgNR8A1 suggested that it might be involved in the embryogenesis of C. gigas. The electrophoretic mobility shift assay showed that CgNR8A1 binds strongly to conserved DNA core motifs DR0, DR2, and DR4 and weakly to DR1, DR3, DR5, Half, and Pal0. In summary, the novel NR8 subfamily identified in this study improves our understanding of NR evolution, and the functional analysis of CgNR8A1 provided further insights into the functions of NR8A1s.