11-P009 The roles of Tbx5 and Tbx4 in limb bud initiation and symmetry

Despite recent progress in understanding of how left–right (L– R) asymmetry is generated during vertebrate development, many important questions still unanswered. One such question concerns the mechanism by which the signal responsible for the generation of L–R asymmetry is transferred from the node to the lateral plate. This signal, whose identity remains unknown, is generated in the node, and its arrival in the left lateral plate induces the asymmetric expression of Nodal. Although it is known that L–R asymmetry-breaking event in the mouse embryo is the leftward fluid flow (nodal flow) on the node, it is unclear what kinds of molecules and signals become asymmetric around the node due to nodal flow. We found that the transcriptional regulatory element, Asymmetric Node Enhancer (ANE), which is located within the 7.5-kb upstream region of human LEFTY-1, could direct left side specific enhancement of reporter gene expression in the node at 1-somite stage, shortly after the beginning of nodal flow. To identify the molecules and signals responsible for the asymmetric activity of ANE, we explored ANE activities in several types of L–R mutant embryos, in which asymmetric Nodal expression in LPM is abnormal. There was a good correlation between the laterality of ANE activity and that of Nodal expression in the LPM. Our results suggest that ANE responds to the L–R asymmetric signals responsible for L–R axis determination. In addition, we want to show how these players interact with each other leading to generate L–R asymmetric signal in the node.