Chikara Meno

Establishment of vertebrate left–right asymmetry

The generation of morphological, such as left–right, asymmetry during development is an integral part of the establishment of a body plan. Until recently, the molecular basis of left–right asymmetry was a mystery, but studies indicate that Nodal and the Lefty proteins, transforming growth factor-β-related molecules, have a central role in generating asymmetric signals. Although the initial mechanism of symmetry breaking remains unknown, developmental biologists are beginning to analyse the pathway that leads to left–right asymmetry establishment and maintenance.

Pitx2. a bicoid-type homeobox gene, is involved in a lefty-signaling pathway in determination of left-right asymmetry

Signaling molecules such as Activin, Sonic hedgehog, Nodal, Lefty, and Vg1 have been found to be involved in determination of left-right (L-R) asymmetry in the chick, mouse, or frog. However, a common signaling pathway has not yet been identified in vertebrates. We report that Pitx2, a bicoid-type homeobox gene expressed asymmetrically in the left lateral plate mesoderm, may be involved in determination of L-R asymmetry in both mouse and chick. Since Pitx2 appears to be downstream of lefty-1 in the mouse pathway, we examined whether mouse Lefty proteins could affect the expression of Pitx2 in the chick. Our results indicate that a common pathway from lefty-1 to Pitx2 likely exists for determination of L-R asymmetry in vertebrates.

Mouse Lefty2 and Zebrafish Antivin Are Feedback Inhibitors of Nodal Signaling during Vertebrate Gastrulation

Mammalian lefty and zebrafish antivin form a subgroup of the TGF beta superfamily. We report that mouse mutants for lefty2 have an expanded primitive streak and form excess mesoderm, a phenotype opposite to that of mutants for the TGF beta gene nodal. Analogously, overexpression of Antivin or Lefty2 in zebrafish embryos blocks head and trunk mesoderm formation, a phenotype identical to that of mutants caused by loss of Nodal signaling. The lefty2 mutant phenotype is partially suppressed by heterozygosity for nodal. Similarly, the effects of Antivin and Lefty2 can be suppressed by overexpression of the nodal-related genes cyclops and squint or the extracellular domain of ActRIIB. Expression of antivin is dependent on Nodal signaling, revealing a feedback loop wherein Nodal signals induce their antagonists Lefty2 and Antivin to restrict Nodal signaling during gastrulation.

Nodal Antagonists in the Anterior Visceral Endoderm Prevent the Formation of Multiple Primitive Streaks

The anterior visceral endoderm plays a pivotal role in establishing anterior-posterior polarity of the mouse embryo, but the molecular nature of the signals required remains to be determined. Here, we demonstrate that Cerberus-like(-/-);Lefty1(-/-) compound mutants can develop a primitive streak ectopically in the embryo. This defect is not rescued in chimeras containing wild-type embryonic, and Cerberus-like(-/-);Lefty1(-/-) extraembryonic, cells but is rescued in Cerberus-like(-/-); Lefty1(-/-) embryos after removal of one copy of the Nodal gene. Our findings provide support for a model whereby Cerberus-like and Lefty1 in the anterior visceral endoderm restrict primitive streak formation to the posterior end of mouse embryos by antagonizing Nodal signaling. Both antagonists are also required for proper patterning of the primitive streak.

Two closely-related left-right asymmetrically expressed genes, lefty-1 and lefty-2: Their distinct expression domains, chromosomal linkage and direct neuralizing activity in Xenopus embryos

Vertebrates have numerous lateral asymmetries in the position of their organs, but the molecular basis for the determination of left–right (L‐R) asymmetries remains largely unknown. TGFβ‐related genes such as lefty and nodal are L‐R asymmetrically expressed in developing mouse embryos, and may be involved in L‐R determination.

Nodal antagonists regulate formation of the anteroposterior axis of the mouse embryo

Patterning of the mouse embryo along the anteroposterior axis during body plan development requires migration of the distal visceral endoderm (DVE) towards the future anterior side by a mechanism that has remained unknown. Here we show that Nodal signalling and the regionalization of its antagonists are required for normal migration of the DVE. Whereas Nodal signalling provides the driving force for DVE migration by stimulating the proliferation of visceral endoderm cells, the antagonists Lefty1 and Cerl determine the direction of migration by asymmetrically inhibiting Nodal activity on the future anterior side.

Planar polarization of node cells determines the rotational axis of node cilia

Rotational movement of the node cilia generates a leftward fluid flow in the mouse embryo because the cilia are posteriorly tilted. However, it is not known how anterior-posterior information is translated into the posterior tilt of the node cilia. Here, we show that the basal body of node cilia is initially positioned centrally but then gradually shifts toward the posterior side of the node cells. Positioning of the basal body and unidirectional flow were found to be impaired in compound mutant mice lacking Dvl genes. Whereas the basal body was normally positioned in the node cells of Wnt3a−/− embryos, inhibition of Rac1, a component of the noncanonical Wnt signalling pathway, impaired the polarized localization of the basal body in wild-type embryos. Dvl2 and Dvl3 proteins were found to be localized to the apical side of the node cells, and their location was polarized to the posterior side of the cells before the posterior positioning of the basal body. These results suggest that posterior positioning of the basal body, which provides the posterior tilt to node cilia, is determined by planar polarization mediated by noncanonical Wnt signalling.

Inhibition of Nodal signalling by Lefty mediated through interaction with common receptors and efficient diffusion

Background: Two TGFβ‐related proteins, Nodal and Lefty, are implicated in early embryonic patterning of vertebrates. Genetic data suggest that Nodal is a signalling molecule, while Lefty is an antagonist of Nodal, but their precise function remains unknown.

GFRα3, a Component of the Artemin Receptor, Is Required for Migration and Survival of the Superior Cervical Ganglion

Abstract GFRα3 is a component of the receptor for the neurotrophic factor artemin. The role of GFRα3 in nervous system development was examined by generating mice in which the Gfrα3 gene was disrupted. The Gfrα3 −/− mice exhibited severe defects in the superior cervical ganglion (SCG), whereas other ganglia appeared normal. SCG precursor cells in the mutant embryos failed to migrate to the correct position, and they subsequently failed to innervate the target organs. In wild-type embryos, Gfrα3 was expressed in migrating SCG precursors, and artemin was expressed in and near the SCG. After birth, SCG neurons in the mutant mice underwent progressive cell death. These observations suggest that GFRα3-mediated signaling is required both for the rostral migration of SCG precursors and for the survival of mature SCG neurons.

Diffusion of Nodal Signaling Activity in the Absence of the Feedback Inhibitor Lefty2

The role of Lefty2 in left-right patterning was investigated by analysis of mutant mice that lack asymmetric expression of lefty2. These animals exhibited various situs defects including left isomerism. The asymmetric expression of nodal was prolonged and the expression of Pitx2 was upregulated in the mutant embryos. The absence of Lefty2 conferred on Nodal the ability to diffuse over a long distance. Thus, Nodal-responsive genes, including Pitx2, that are normally expressed on the left side were expressed bilaterally in the mutant embryos, even though nodal expression was confined to the left side. These results suggest that Nodal is a long-range signaling molecule but that its range of action is normally limited by the feedback inhibitor Lefty2.