Characterization of NF-kappa B/I kappa B proteins in zebra fish and their involvement in notochord development

Although largely involved in innate and adaptive immunity, NF-kappa B plays an important role in vertebrate development. In chicks, the inactivation of the NF-kappa B pathway induces functional alterations of the apical ectodermal ridge, which mediates limb outgrowth. In mice, the complete absence of NF-kappa B activity leads to prenatal death and neural tube defects. Here, we report the cloning and characterization of NF-kappa B/I kappa B proteins in zebra fish. Despite being ubiquitously expressed among the embryonic tissues, NF-kappa B/I kappa B members present distinct patterns of gene expression during the early zebra fish development. Biochemical assays indicate that zebra fish NF-kappa B proteins are able to bind consensus DNA-binding (kappa B) sites and inhibitory I kappa B alpha proteins from mammals. We show that zebra fish I kappa B alphas are degraded in a time-dependent manner after induction of transduced murine embryo fibroblasts (MEFs) and that these proteins are able to rescue NF-kappa B activity in I kappa B alpha(-/-) MEFs. Expression of a dominant-negative form of the murine I kappa B alpha (mI kappa B alpha M), which is able to block NF-kappa B in zebra fish cells, interferes with the notochord differentiation, generating no tail (ntl)-like embryos. This phenotype can be rescued by coinjection of the T-box gene ntl (Brachyury homologue), which is typically required for the formation of posterior mesoderm and axial development, suggesting that ntl lies downstream of NF-kappa B . We further show that ntl and Brachyury promoter regions contain functional kappa B sites and NF-kappa B can directly modulate ntl expression. Our study illustrates the conservation and compatibility of NF-kappa B/I kappa B proteins among vertebrates and the importance of NF-kappa B pathway in mesoderm formation during early embryogenesis.