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In Xenopus eggs, fertilization initiates a rotational movement of the cortex relative to the cytoplasm, resulting in the transport of critical determinants to the future dorsal side of the embryo. Cortical rotation is mediated by microtubules, resulting in activation of the Wnt/β-catenin signaling pathway and expression of organizer genes on the dorsal side of the blastula. Similar cytoplasmic localizations resulting in β-catenin activation occur in many chordate embryos, suggesting a deeply conserved mechanism for patterning early embryos. This review summarizes the experimental evidence for the molecular basis of this model, focusing on recent maternal loss-of-function studies that shed light on two main unanswered questions: (1) what regulates microtubule assembly during cortical rotation and (2) how is Wnt/β-catenin signaling activated dorsally? In addition, as these processes depend on vegetally localized molecules in the oocyte, the mechanisms of RNA localization and novel roles for localized RNAs in axis formation are discussed. The work reviewed here provides a beginning framework for understanding the coupling of asymmetry in oogenesis with the establishment of asymmetry in the embryo.
Figure 1. Conceptual model for dorsal axis formation in Xenopus. (a) Oogenesis. During oogenesis, β-catenin-stabilizing factors are localized to the vegetal pole. Dorsoventral polarity is not apparent at this stage, and the oocyte and mature egg exhibit axisymmetry about the animal–vegetal axis. (b) Cortical rotation. Following fertilization, the cortex of the egg rotates relative to the inner cytoplasm using a parallel array of microtubules. Movement generally occurs opposite to the sperm entry point (SEP), and transports cortical and subcortical β-catenin-stabilizing dorsalizing activity into the equatorial region of the embryo. (c) β-Catenin stabilization/nuclear localization. During the 16–128-cell stage, β-catenin is stabilized and is enriched in dorsal nuclei, remaining nuclear through the midblastula transition (MBT). (d) Organizer gene expression. At the late blastula/early gastrula stage, β-catenin (along with other proteins, such as Vegt) activates gene expression in the dorsal vegetal and equatorial regions. These targets include Spemann organizer genes siamois, nodal-related 3, chordin, and noggin, as well as early vegetal Nodal homologs, nodal-related 5/6. These and other genes regulate axial patterning by antagonizing the function of bone morphogenetic proteins as well as regulating other signaling molecules. D, dorsal side; GV, germinal vesicle; V, ventral side.
Figure 2. Cellular events of cortical rotation. (a and b) Assembly and alignment of microtubules. (a) Thirty to forty minutes after fertilization. Astral microtubules organized around the spermcentriole and pronucleus grow radially. Microtubules also originate in the cortex/subcortical region and within the inner cytoplasm. Kinesins and related proteins are enriched in the vegetal cortex. Plus ends are indicated by arrows; the relative scale of the shear zone is exaggerated for clarity. (b) Sixty to eighty minutes after fertilization. The egg undergoes cortical rotation. Microtubules in the shear zone are aligned in parallel and oriented toward the future dorsal side. (c and d) Models showing the displacement of the cortex relative to the inner cytoplasm. Only the vegetal cortex is shown (light blue); the region of dorsalizing activity is shaded dark blue. The heavy dashed line indicates the initial animal–vegetal axis; the smaller dashed line indicates the position of the former vegetal pole. (e and f) Immunostaining against α-tubulin. Vegetal views of eggs at 50 min postfertilization (e) and at 80 min postfertilization (f), showing parallel alignment and bundling of microtubules.
Figure 3. Models for Wnt/β-catenin signaling regulation during Xenopus axis formation. (a) Diagram of overall Wnt/β-catenin signaling. In ventral cell, β-catenin is phosphorylated by the destruction complex [Axin/adenomatous polyposis coli (APC)/GSK3β] and targeted for degradation. In those nuclei, Tcf3 represses critical dorsal gene expression. In a dorsal cell, Wnt activation leads to inhibition of destruction complex activity and accumulation of free β-catenin, which can enter the nucleus. β-Catenin is assembled into transcriptional complexes with Tcf3/Pygo/Legless at sites primed by Prmt2 and dorsal genes are expressed. (b) Potential mechanisms of β-catenin activation in the Xenopus embryo. (1) Enrichment of wnt11 RNA dorsally, followed by activation by secreted Wnt11/5a/Frl1 complexes or (2) an unknown Wnt ligand; (3) intracellular activation by Frat1/Dvl2 particles or (4) an unknown component of vegetal cortical cytoplasm (VCC). Predominantly positive-acting components are shown in green, negative components in red. Molecules with potentially dual or noncanonical roles are shown in yellow. Arrowed lines indicate positive regulation and ‘T’ bars indicate negative regulation. The arrangement of the boxes is not meant to convey specific exact binding relationships or stoichiometry.
Figure 4. Localized messenger RNA (mRNA) in Xenopus. (a) Expression of a germ plasm-localized mRNA during oogenesis, showing restriction to the mitochondrial cloud (m.c.; arrow) in stage I oocytes. Expression in later stages (stage III–VI) is localized to a compact domain at the vegetal apex. (b) Expression of a late pathway mRNA in a stage VI oocyte (vegetal view). Expression is broadly distributed throughout the vegetal hemisphere. Inset shows nonlocalized expression in a stage I oocyte. Images in (a) and (b) are not to scale. (c) Diagram of mRNA localization pathway in Xenopus oocytes. Germ plasm-localized mRNAs are indicated in dark blue, late pathway mRNAs are in light blue. Microtubules involved in late localization are in green. GV, germinal vesicle. Oocyte stages are indicated above each diagram.
