Ascidians, along with other urochordates, are the most evolutionary distant group from vertebrates to display definitive chordate-specific characters, such as a notochord, dorsal hollow nerve cord, pharynx and endostyle. Most solitary ascidians have a biphasic life history that has partitioned the development of these characters between a planktonic microscopic tadpole larva (notochord and dorsal nerve cord) and a larger sessile adult (pharynx and endostyle). Very little is known of the molecular axial patterning processes operating during ascidian postlarval development. Two axial patterning homeobox genes Otx and Cdx are expressed in a spatially restricted manner along the ascidian anteroposterior axis during embryogenesis and postlarval development (i.e., metamorphosis). Comparisons of these patterns with those of homologous cephalochordate and vertebrate genes suggest that the novel ascidian biphasic body plan was not accompanied by a deployment of these genes into new pathways but by a heterochronic shift in tissue-specific expression. Studies examining the role of all-trans retinoic acid (RA) in axial patterning in chordates also contribute to our understanding of the role of homeobox genes in the development of larval and adult ascidian body plans. Our studies demonstrate that RA does not regulate axial patterning in the developing ascidian larval neuroaxis in a manner homologous to that found in vertebrates. Although RA may regulate the expression of some ascidian homeobox genes, ectopic application of RA does not appear to alter the morphology of the larval CNS. However, treatment with similar or lower concentrations of RA, have a profound effect on postlarval development and the juvenile body plan. These changes are correlated to a dramatic reduction of Otx expression. Through these RA-induced effects we infer that while RA may regulate the expression of some homeobox genes during embryogenesis it has a far more dramatic impact on postlarval development where regulative processes predominate.