Early control of left-right patterning and bilateral symmetry in Xenopus embryos

MONTI, RENATO JOSÉ; REVINSKI, DIEGO R.; TICONA OQUENDO, RUBÉN M.; CASTRO COLABIANCHI, AITANA M.; GONZÁLEZ STIVALA, ERNESTO; LÓPEZ, SILVIA L.

Santos, São Paulo

Congreso; 8th Meeting of the Latin American Society of Developmental Biology (LASDB); 2015

Latino American Society for Developmental Biology

Although the body of vertebrates display a bilateral external appearance, the morphology, location andorientation of several internal organs such as the intestine and the heart show a consistent left-right (LR) asymmetry which is evolutionarily conserved in amphibians, reptiles, birds, fishes and mammals. When LR patterning fails, congenital disorders like mild to severe heterotaxy arise, which sometimes are lethal. By performing functional experiments in Xenopus laevis, we have previously shown that an early ventralNotch activity contribute to restrict the BCNE center to the dorsal side of the blastula (Acosta et al. 2011, Development 138, 2567?79). Now, we present evidence that the Notch protein is asymmetrically distributed through the dorsal-ventral axis (confirming our previous hypothesis) and noteworthy, also does across the left-right axis from the earliest stages of development, with higher levels on the left side. Since Notch is a well known substrate of γ-secretase, we performed temporally controlled experiments by treating embryos with the inhibitor DAPT during different time windows. We found that treatments during cleavage stages not only produced different grades of heterotaxy but, surprisingly, also resulted in asymmetries of normally bilateral symmetric structures of the body. To ascertain if Notch signaling is involved in early mechanisms controlling LR patterning and bilateral symmetry, we began working with antisense Photo-Mopholinos against Notch. We injected albino embryos and irradiated them with UV light at blastula stage in order to circumscribe inhibition of Notch signaling to early cleavage stages. N-tubulin expression in neurulae indicated that the Photo-Mo effectively blocked Notch signaling and that this effect could be reverted by UV irradiation. Moreover, the alterations of LR patterning and bilateral symmetry that we observed after UV irradiation of Photo-MO injected embryos suggest that a left bias of Notch signaling is established during early development. In overall, our results indicate that: 1) the bilateral symmetry of normally symmetric structures is not a default stage and requires early intact γ-secretase and Notch activities; 2) Notch signaling regulates LR patterning much earlier than previously acknowledged.