Determination of the vertebrate left-right body axis during embryogenesis results in asymmetric development and placement of most inner organs. Although the asymmetric Nodal cascade is conserved in all vertebrates, the mechanism of symmetry breakage has remained controversial. In mammalian and fish embryos, a cilia-driven leftward flow of extracellular fluid is required for initiation of the Nodal cascade. This flow is localized at the posterior notochord ("node") and Kupffer's vesicle, respectively. In frog and chick embryos, however, molecular asymmetries are required earlier, from cleavage stages through gastrulation. The validity of a cilia-based mechanism for all vertebrates therefore has been questioned. Here we show that a cilia-driven leftward flow precedes asymmetric nodal expression in the frog Xenopus. Motile monocilia emerged on the gastrocoel roof plate during neurulation and lengthened and polarized from an initially central position to the posterior pole of cells. Concomitantly, a robust leftward fluid flow developed from stage 15 onward, significantly before asymmetric nodal transcription started in the left-lateral-plate mesoderm at stage 19. Injection of 1.5% methylcellulose into the archenteron prevented leftward flow and resulted in laterality defects, demonstrating that the flow itself was required for asymmetric gene expression and organ placement.
In vertebrates, the readily apparent left-right (L/R) anatomical asymmetries of the internal organs can be traced to molecular events initiated at or near the time of gastrulation. However, the earliest steps of this process do not seem to be universally conserved. In particular, how this axis is first defined in chicks has remained problematic. Here we show that asymmetric cell rearrangements take place within chick embryos, creating a leftward movement of cells around the node. It is the relative displacement of cells expressing Sonic hedgehog (Shh) and Fibroblast growth factor 8 (Fgf8) that is responsible for establishing their asymmetric expression patterns. The creation of asymmetric expression domains as a passive effect of cell movements represents an alternative strategy for breaking L/R symmetry in gene activity.In mice and rabbits monocilia responsible found on cells of the posterior notochordal plate have been shown to play a crucial role in breaking L/R symmetry (1,2). These cilia are able to create a leftward flow of fluid, in a pit-like teardrop shaped space that is not covered by subjacent endoderm (3)., The flow of fluid across this pit stimulates signal transduction that ultimately leads to induction of asymmetric gene expression (1,2).In the chick embryo, in contrast, the endoderm underlying Hensen's node (a structure at the rostral end of the primitive streak in the gastrulating embryo) exists as a continuous sheet ventral to the notochordal plate mesoderm (4) and there is no morphological pit on the ventral surface in which a flow of fluid could be established. Prior work has noted cilia at Hensen's node (5) but these short cilia are on endodermal cells and are unrelated to the motile cilia on the mesodermal cells of the ventral node in the mouse and rabbit. The mesodermal cells at Hensen's node in the chick are devoid of cilia. In addition, the Talpid chick mutant lacks primary cilia (6) but does not exhibit L/R asymmetry defects. Unlike the mouse and rabbit, the chick node itself becomes morphologically asymmetric, with a marked tilt towards the left around the time the primitive streak reaches full extension, at Stage 4. (7,8) (Fig. 1A-C). Shortly thereafter, a number of small L/R asymmetric expression domains are observed to the right and left of the node (9). However, all of the genes expressed in such a manner are initially expressed in a symmetric fashion, for example, Fgf8 bilaterally along the primitive streak and Shh bilaterally across the top of the node until stage 4, (10) (Fig. 1D_G) NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript morphological asymmetries in the node, these gene expression patterns also become gradually asymmetric by stage 5 (Fig. 1H,I).To investigate cellular rearrangements that could be responsible for establishing the morphological asymmetry of the node, we performed a time lapse analysis of cell movements at Hensen's node; randomly labeling cells by electroporation of a green fluorescent protein (GFP) reporter. At stage 4, ...
Morphological asymmetry is a common feature of animal body plans, from shell coiling in snails to organ placement in humans. The signaling protein Nodal is key for determining this laterality. Many vertebrates, including humans, use cilia for breaking symmetry during embryonic development: rotating cilia produce a leftward flow of extracellular fluids that induces the asymmetric expression of Nodal. By contrast, Nodal asymmetry can be induced flow-independently in invertebrates. Here, we ask when and why flow evolved. We propose that flow was present at the base of the deuterostomes and that it is required to maintain organ asymmetry in otherwise perfectly bilaterally symmetrical vertebrates.
Motile monocilia play a pivotal role in left-right axis determination in mouse and zebrafish embryos. Cilia with 9؉0 axonemes localize to the distal indentation of the mouse egg cylinder ("node"), while Kupffer's vesicle cilia in zebrafish show 9؉2 arrangements. Here we studied cilia in a prototype mammalian embryo, the rabbit, which develops via a flat blastodisc. Transcription of ciliary marker genes
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