The Role of Brachyury (T) during Gastrulation Movements in the Sea Urchin Lytechinus variegatus

Abstract

The studies described here sought to identify and characterize genes involved in the gastrulation and morphogenetic movements that occur during sea urchin embryogenesis. An orthologue of the T-box family transcription factor, Brachyury, was cloned through a candidate gene approach. Brachyury (T) is the founding member of this T-box transcription factor family and has been implicated in gastrulation movements in Xenopus, zebrafish, and mouse embryogenesis. Polyclonal serum was generated to LvBrac in order to characterize protein expression. LvBrac initially appears at mesenchyme blastula stage in two distinct regions with embryonic expression perduring until pluteus stage. Vegetally, LvBrac expression is in endoderm and lies circumferentially around the blastopore. This torus-shaped area of LvBrac expression remains constant in size as endoderm cells express LvBrac upon moving into that circumference and cease LvBrac expression as they leave the circumference. Vegetal expression remains around the anus through pluteus stage. The second domain of LvBrac expression first appears broadly in the oral ectoderm at mesenchyme blastula stage and at later embryonic stages is refined to just the stomodael opening. Vegetal LvBrac expression depends on autonomous β-catenin signaling in macromeres and does not require micromere or veg2-inductive signals. It was then determined that LvBrac is necessary for the morphogenetic movements occurring in both expression regions. A dominant-interfering construct was generated by fusing the DNA binding domain of LvBrac to the transcriptional repression module of the Drosophila Engrailed gene in order to perturb gene function. Microinjection of mRNA encoding this LvBrac-EN construct resulted in a block in gastrulation movements but not expression of endoderm and mesoderm marker genes. Furthermore, injection of LvBrac-EN into one of two blastomeres resulted in normal gastrulation movements of tissues derived from the injected blastomere, indicating that LvBrac downstream function may be nonautonomous during sea urchin gastrulation.