Expression of GD3 Synthase and paraxis transcription factor in mesodermal layer formation during Xenopus laevis embryogenesis.

LUQUE M.E.; MÓNACO M.E.; SANCHEZ S.S.

Pinamar. Buenos Aires. Argentina

Congreso; X Congress of the Panamerican Association for Biochemistry and Molecular Biology (PABMB), the XLI Annual Meeting of the Argentine Society for Biochemistry and Molecul; 2005

Association for Biochemistry and Molecular Biology (PABMB), the Argentine Society for Biochemistry and Moleculecular Biology

Glycosphingolipids (GSL) are believed to be integral for the dynamics of many cell membrane events, including cellular interactions, signaling and trafficking. In previous studies we investigated the ganglioside profile, cellular expression, and biosynthesis during the early development of the Xenopus laevis amphibian embryo. In this work we report the molecular cloning of a2,8-sialyltrasnsferase (GD3 synthase, GD3s), a key enzyme that catalizes the first step of b-series gangliosides synthesis. We have isolated a 2.88 Kb length transcript for GD3s. Expression of this enzyme was analyzed during early development of Xenopus embryos by RT-PCR and in situ hybridization. We detected transcripts of GD3s from stage 10 onward. The spatio-temporal analysis of expression patterns of enzyme and specific markers by in situ hybridization reveals that it is developmentally regulated. GD3 is mainly expressed in dorsal blastopore lip at gastrula stages and in the migrating mesodermal cells. In later stages of development it is expressed in the presomitic mesoderm together with the paraxis transcription factor. Experiments using a resin microsphere embedded with PPMP (phenyl palmitoyl morpholino propanol), a specific inhibitor of glycosphingolipid byosinthesis, was implanted in the right side of stage 10 embryos. The inhibitor produced embryos with abnormal expression of mesodermal markers, such as goosecoid, chordin. Knock-down experiments using expression of a dominant negative form paraxis revealed severe defects in somite formation. Rescue experiments confirmed the specificity of this experiment. We are currently investigating a possible role of paraxis transcription factor in mesodermal formation. Together, our results strongly argue for a role of gangliosides in the formation presomitic area and raised the questions if paraxis is implicated