Initiating the Embryonic Body Plan: Dorsalization of the Xenopus Embryo
What is the role of the wingless pathway in dorsalization?
The evidence is strong that dorsal determinants exist in the Xenopus embryo. The
identification of dorsal determinants is an important goal for understanding how the body
plan is established. Valuable clues to the identities of the factors that dorsalize the Xenopus
embryo came from experiments demonstrating that injections ofWnt mRNA will cause
duplication of the Xenopus embryonic axis (McMahon and Moon, 1989; Smith and
Harland, 1991; Sokol et al., 1991; see Fig.1). The Wnt proteins are a family of
growth factors that are homologues of the wingless segment polarity gene of Drosophila.,
which maintains the expression of the engrailed gene, which encodes a transcription
factor. The wingless signaling pathway has been described in Drosophila.
Therefore, attempts have been made to examine the possible roles of the Wnts and
members of the Wnt signal transduction pathway in dorsal-ventral axis specification
wingless -> frizzled ->dishevelled -| zw3/shaggy -| armadillo -> cell fate
frizzled encodes the wg receptor, whereas the remaining factors are intracellular. armadillo , which interprets the wingless signal, encodes a protein that associates with the cytoplasmic domains of members of the cadherin family of cell adhesion proteins. Interestingly, there is considerable recent evidence that the cadherins may initiate signal transduction cascades in addition to their roles in adhesion.
Vertebrate homologues of the protein encoded byarmadillo include ß-catenin and plakoglobin. The protein that is immediately upstream of armadillo (zw3/shaggy) is a serine/threonine kinase. Wingless signaling causes a decrease in the phosphorylation of the armadillo protein and an increase in its stablility. Thus, zw3/shaggy is thought to regulate armadillo by phosphorylation.
Xenopus eggs contain transcripts encoding components of the Wnt pathway.We have previously discussed the accumulation and localization of Xwnt-11 transcripts during oogenesis. Other Wnt homologs in the egg include Xwnt-5A and Xwnt-8b. Transcripts of a dishevelled homolog, a zw3/shaggy homolog (Xgsk-3), an armadillo homolog and ß-catenin. have also been demonstrated (for review, see Yost, Torres et al., 1996). Do they play any role in axis specification, and - if so - what might it be?
We have already discussed the effects of overexpression of Wnt; it causes axis duplication. Overexpression of other components of the Wg signal transduction cascade does the same. An example is shown below, which shows neural axis duplication after injection of plakoglobin RNA (from Mike Klymowsky; copyright ©, 1996, Mike Klymkowsky; Karnovsky and Klymkowsky, 1995).
Strong evidence that the Wnt pathway is required for axis determination was provided by Heasman et al. (1994), who demonstrated that depletion of oogenic ß-catenin transcripts using antisense oligo technology caused reductions in dorsal structures, an effect that could be rescued by injection of ß-catenin mRNA (Figs. 3-5). Depletion of ß-catenin could not be rescued by ectopic Wnt signals, a result that is consistent with ß-catenin being downstream of Wnt.
How are the effects of ß-catenin/plakoglobin produced? Heasman et al. demonstrated that one defect of ß-catenin depletion is the failure to produce dorsal mesoderm (the organizer). As we shall discuss later, the organizer is induced by the underlying Nieuwkoop center. This suggests that ß-catenin is required for the signaling events that are necessary for dorsal mesoderm induction by the Nieuwkoop center.
Randall Moon has prepared an overview of the role of ß-catenin in Xenopus development. Study his overview and then return here.
As discussed by Dr. Moon, the target of ß-catenin/plakoglobin may be a transcription factor called XTcf-3 (Molenaar et al., 1996). XTcf-3 is a homolog of a group of vertebrate high mobility group (HMG) box transcription factor genes. Its mammalian counterparts are referred to as "architectural transcription factors" that affect spatial structure of enhancers of target genes, thus facilitating contacts between other factors that are bound to the enhancer. Transcripts encoding XTcf-3 are present in Xenopus unfertilized eggs and early embryos. These transcripts are most abundant in the animal hemisphere and in the marginal zone of the embryo (Molenaar et al., 1996, Fig. 1). In vitro assays demonstrated that XTcf-3 is capable of binding to ß-catenin. XTcf-3 and ß-catenin also interact in vivo, which causes the translocation of ß-catenin into the nucleus (Fig. 4). An N-terminal deletion mutant of XTcf-3 (delta N) that is incapable of interacting with ß-catenin failed to translocate ß-catenin into the nucleus. This mutant acts as a dominant-negative mutant that presumably perturbs the normal interaction between ß-catenin and endogenous XTcf-3 (Molenaar et al., 1996).
If delta N perturbs ß-catenin function, one would predict that it would prevent the ability of overexpressed ß-catenin RNA to induce axis duplication. As shown in Table 1 and Figure 6, this prediction has been upheld. Would it also perturb the function of endogenous ß-catenin? (Remember that depletion of endogenous ß-catenin will suppress axis formation.) Indeed, axis formation is perturbed when RNA is injected into the two dorsal blastomeres at the 4-cell stage, and the effect is most effective when the injections of the RNA encoding the dominant-negative XTcf-3 mutant were done in the marginal zone (Table 2 and Fig. 7).
Hedgepeth, C.M., Conrad, L.J., Zhang, J., Huang, H.-C., Lee, V.M.Y. and Klein, P.S. 1997. Activation of the Wnt signaling pathway: a molecular mechanism for lithium action. Develop. Biol. 185: 82-91.
Kageura, H. 1997. Activation of dorsal development by contact between the cortical dorsal determinant and the equatorial core cytoplasm in eggs of Xenopus laevis. Development 124: 1543-1551.
Kofron, M., Spagnuolo, A., Klymkowsky, M., Wylie, C. and Heasman, J. 1997. The roles of maternal alpha-catenin and plakoglobin in the early Xenopus embryo. Development 124: 1553-1560.
Merriam, J.M., Rubenstein, A.B. and Klymkowsky, M.W. 1997. Cytoplasmically anchored plakoglobin induces a WNT-like phenotype in Xenopus. Develop. Biol. 185: 67-81.
Miller, J.R., Rowning, B.A., Larabell, C.A., Yang-Snyder, J.A., Bates, R.L., and Moon, R.T. 1999. Establishment of the dorsal-ventral axis in Xenopus embryos coincedes with the dorsal enrichment of dishevelled that is dependent on cortical rotation. J. Cell. Biol. 146:427-437.
Wodarz, A. and Nusse, R. 1998. Mechanisms of Wnt signaling in development. Annu. Rev. Cell Dev. Biol. 14:59-88.
Links to Related Material
Evidence that dorsal determinants exist in the Xenopus embryo.
The accumulation and localization of Xwnt-11 transcripts during oogenesis.
See the recent cautionary note in BioEssays by Mike Klymkowsky.
See "Transduction of the Wingless Signal to the Nucleus" in Zygote.
Browder, L.W., C. A. Erickson and W.R. Jeffery. 1991. Developmental Biology.
Third Edition. Saunders College Publishing. Philadelphia.
Molenaar, M. et al. 1996. XTcf-3 transcription factor mediates ß-catenin-induced axis formation in Xenopus embryo. Cell 86: 391-399.
Smith, W.C. and Harland, R.M. 1991. Injected Xwnt-8 RNA acts early in Xenopus embryos
to promote formation of a vegetal dorsalizing center. Cell 67: 753-765.
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Leon Browder & Laurie Iten (Ed.) Dynamic Development
Last revised Friday, November 6, 1998