Synergistic action of HNF-3 and Brachyury in the notochord differentiationof ascidian embryos

In vertebrate embryos, the class I subtype forkhead domain gene HNF-3 is es sential for the formation of the endoderm, notochord and overlying ventral neural tube. In ascidian embryos, Brachyury is involved in the formation of the notochord. Although the results of previous studies imply a role of HN F-3 in notochord differentiation in ascidian embryos, no experiments have b een carried out to address this issue directly. Therefore the present study examined the developmental role of HNF-3 in ascidian notochord differentia tion. When embryos were injected with a low dose of HNF-3 mRNA, their tails were shortened and when embryos were injected with a high dose of HNF-3 mR NA, which was enough to inhibit differentiation of epidermis and muscle, no obvious ectopic differentiation of endoderm or notochord cells was observe d. However, co-injection of HNF-3 mRNA along with Brachyury mRNA resulted i n ectopic differentiation of notochord cells in the animal hemisphere, sugg esting that HNF-3 acts synergistically with Brachyury in ascidian notochord differentiation. Notochord differentiation of the A-line precursor cells d epends on inducing signal(s) from endodermal cells, which can be mimicked b y bFGF treatment. Treatment of notochord precursor cells isolated from the 32-cell stage embryos with bFGF resulted in upregulation of both the HNF-3 and Brachyury genes.