Ci-FoxA-a is the earliest zygotic determinant of the ascidian anterior ectoderm and directly activates Ci-sFRP1/5

This work focuses on the anteroposterior patterning of the ectoderm in the invertebrate chordate Ciona intestinalis. Previous work indicated that, by the eight-cell stage, the anterior and posterior animal blastomeres have acquired different properties, including a differential responsiveness to inducing signals from the underlying mesendoderm. Here, we investigated the molecular basis of this distinction. For this, we studied the regulation of the earliest marker specific for the anterior ectoderm, Ci-sFRP1/5, which is activated at the 64-cell stage. We first found that the activation of this marker in the anterior ectoderm does not involve communication with other lineages. We then identified, by phylogenetic footprinting and deletion analysis, a short conserved minimal enhancer driving the onset of expression of Ci-sFRP1/5. We showed that this enhancer was a direct target of the Ci-FoxA-a gene, a FoxA/HNF3 orthologue expressed in anterior ectodermal and mesendodermal lineages from the eight-cell stage. Gain-and loss-of-function experiments revealed that Ci-FoxA-a is necessary and sufficient within the ectoderm to impose an ectodermal anterior identity, and to repress the posterior programme. Thus, Ci-FoxA-a constitutes a major early zygotic anterior determinant for the ascidian ectoderm, acting autonomously in this territory, prior to the onset of vegetal inductions. Interestingly, while vertebrate FoxA2 are also involved in the regionalization of the ectoderm, they are thought to act during gastrulation to control, in the mesendoderm, the expression of organizer signals. We discuss the evolution of chordate ectodermal patterning in light of our findings.