1 Division of Morphogenesis, Department of Developmental Biology, National
Institute for Basic Biology, 38 Nishigonaka, Myodaiji, Okazaki 444-8585,
Japan
2 Department of Anatomy and Developmental Biology, University College London,
Gower Street, London WC1E 6BT, UK
* Author for correspondence (e-mail: nueno{at}nibb.ac.jp)
Accepted 12 February 2003
![]() |
SUMMARY |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: Heparan sulfate proteoglycan, Wnt signaling pathway, Gastrulation movements, Xenopus
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Recently, it has been reported that a non-canonical Wnt signaling cascade,
which is known to regulate planar cell polarity (PCP) in Drosophila
(Adler, 1992;
Mlodzik, 1999
), also
participates in the regulation of convergent extension movements in
Xenopus, as well as in the zebrafish embryo
(Djiane et al., 2000
;
Heisenberg et al., 2000
;
Tada and Smith, 2000
;
Wallingford et al., 2000
). The
zebrafish silberblick (slb) locus encodes Wnt11, and
Slb/Wnt11 activity is required for cells to undergo correct convergent
extension movements during gastrulation. The signal transducer Dishevelled
(Dsh) acts downstream of Slb/Wnt11 through domains specific to the
non-canonical Wnt/PCP signaling cascade, and directly regulates cell polarity
within cells undergoing convergent extension
(Heisenberg et al., 2000
;
Tada and Smith, 2000
;
Wallingford et al., 2000
). In
addition, the relocalization of Dsh to the cell membrane is required for
convergent extension movements in Xenopus gastrulae, in the same way
as recruitment of Dsh to the membrane through the Frizzled (Fz) receptor is
required for the PCP pathway in Drosophila
(Axelrod, 2001
;
Wallingford et al., 2000
).
Although many genetic experiments have demonstrated downstream effectors of
the non-canonical PCP Fz/Dsh signal in Drosophila
(Shulman et al., 1998
;
Tree et al., 2002
), and some
of these effectors (e.g. JNK) function to regulate the convergent extension
movements (Darken et al.,
2002
; Yamanaka et al.,
2002
), the precise signaling mechanism that regulates the
convergent extension movements remains unclear.
In addition to the intracellular signaling mechanism, intercellular
modulators are involved in regulating coordinated movements of large cell
populations. Heparan sulfate proteoglycans (HSPGs) have been implicated in the
modulation of intercellular signaling in vertebrates and in
Drosophila, and have been shown to be required for gastrulation
movements in the Xenopus embryo
(Brickman and Gerhart, 1994;
Itoh and Sokol, 1994
).
Glypican, a member of the membrane-associated HSPG family, is known to
regulate Wnt signaling in Drosophila
(Baeg et al., 2001
;
Tsuda et al., 1999
) and
zebrafish embryos (Topczewski et al.,
2001
). However, the molecular function of the HSPGs is poorly
understood. Therefore, we cloned a gene encoding Xenopus glypican 4
(Xgly4), a member of the HSPG family, and analyzed its role in the
non-canonical Wnt/PCP signaling pathway during gastrulation. In situ
hybridization revealed that Xgly4 is expressed in the dorsal mesoderm
and ectoderm during gastrulation. Xgly4 overexpression and translational
inhibition by a morpholino oligonucleotide inhibited the gastrulation
movements of the embryo and the convergent extension (elongation) of
activin-treated animal caps, but did not affect mesoderm induction. Rescue
analysis with different Dsh mutants and Wnt11 demonstrated that Xgly4
functions in the non-canonical Wnt/PCP pathway, but not in the canonical
Wnt/ß-catenin pathway for convergent extension movements. Furthermore, we
show that reducing the levels of Xgly4 inhibits the cell-membrane accumulation
of Dsh, as does the inhibition of Wnt11 in the dorsal marginal zone. Finally,
we provide evidence that the Xgly4 protein physically binds Wnt ligands.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The Xgly4 morpholino sequence was
5'-TGCATGGTGAGAACGAAAGCGATC-3'. We used the morpholino
oligonucleotide 5'-CATAGATGTATAAACGCTACGAAAG-3' for the ascidian
gene HrTT-1 (Hotta et al.,
1998) as a negative control (Gene Tools, LLC)
Manipulation of embryos, microinjection of synthetic mRNA and
morpholino oligonucleotide
Unfertilized eggs were collected and fertilized in vitro as described
previously (Suzuki et al.,
1997). Embryos were de-jellied using 3% cysteine and washed with
water several times. Embryos were staged according to Nieuwkoop and Faber
(Nieuwkoop and Faber, 1967
).
Synthesized RNA was injected into the animal pole or marginal zone of two- or
four-cell-stage embryos, respectively, and the injected embryos were kept in
3% Ficoll/0.1x Steinberg's solution as described
(Yamamoto et al., 2000
). The
animal cap (AC) and dorsal marginal zone (DMZ) were dissected at stage 8 and
stage 10.5, respectively, and cultured in 0.1% BSA/1x Steinberg's
solution (Asashima et al.,
1990
). For the standard Xenopus AC assay, ACs were
cultured until stage 18, and scored for elongation.
In situ hybridization
Whole-mount in situ hybridization was performed with digoxigenin
(DIG)-labeled probes, as described by Harland
(Harland, 1991). Hybridization
was detected with an alkaline phosphatase-coupled anti-DIG antibody and
visualized by using BM purple (Roche Molecular Biochemicals). Antisense in
situ probes against Xgly4 and Xwnt11 were generated by linearizing the
pBS-KS-Xgly4 and pBS-KS-Xwnt11 constructs with NotI, and transcribing
them with T7 RNA polymerase. Xbra (Smith
et al., 1991
), XmyoD (Hopwood
et al., 1989
), Xgsc (Blumberg
et al., 1991
; Cho et al.,
1991
), Xvent1 (Nishimatsu and
Thomsen, 1998
) and Xnot (von
Dassow et al., 1993
) have been reported previously.
RT-PCR analysis
Profiles of Xbra and Xwnt11 expression in the animal cap
injected with each mRNA or morpholino oligonucleotide were examined
semi-quantitatively using RT-PCR (Yamamoto
et al., 2000). ACs were collected when sibling embryos had
developed to stage 11. The total RNA was then isolated using TRIzol
(GIBCO/BRL), and cDNA was synthesized as previously reported
(Yamamoto et al., 2000
). PCR
was performed with oligonucleotide primers: Xbra,
5'-GGATCATCTTCTCAGCGCTGTGGA-3' (upsteam) and
5'-GTTGTCGGCTGCCACAAAGTCCA-3' (downstream); and Xwnt11,
5'-AAGTGCCACGGAGTGTCTGG-3' (upstream) and
5'-CTCAGACTCTCTCACTGGCC-3' (downstream). The primer sequence of
histone, an internal input control, was as previously described
(Iemura et al., 1998
).
Northern blot analysis
PolyA+ RNA was extracted from whole oocytes and staged embryos
using TRIzol reagent (GIBCO/BRL) and oligotex (Roche). Oocytes and embryos
were staged according to Nieuwkoop and Faber
(Nieuwkoop and Faber, 1967).
RNA was run on a 1% agarose gel containing formaldehyde, blotted onto nylon
membranes, and hybridized with a DNA probe in 50% formamide, 5xSSPE, 5%
SDS and 100 µg/ml denatured salmon sperm DNA. The entire Xgly4 cDNA was
used as the probe. Blots were hybridization overnight at 55°C and then
washed twice in 0.1xSSPE, 0.1% SDS at 55°C for 1 hour each.
Immunofluorescence
The DMZ and AC were dissected, fixed in MEMFA and incubated with anti-Myc
antibody (9E10, Santa Cruz Biotechnology; 1:200) for 16 hours at 4°C.
Subsequently, these tissues were incubated with an FITC-labeled secondary
antibodies (anti-mouse IgG goat serum, 1:200) for 2 hours at room temperature,
and examined using fluorescence microscopy
(Larabell et al., 1997). To
quantitate the localization of Xdsh in cells, five cells were randomly chosen
from the mRNA- or morpholino oligonucleotide-injected area, which was marked
by fluorescence, and the intensity of Xdsh staining in the cytoplasm and
membrane in each cell was quantitated by using NIH image. The closest four
among five sets of data were processed for graphical presentation.
Immunoprecipitation
HEK293T cells were transiently transfected with the indicated constructs
using the calcium phosphate method. Forty hours after transfection, the cells
were lyzed in lysis buffer [50 mM Tris-HCl (pH 7.5), 150 mM NaCl, 5 mM EDTA,
0.5% NP-40, 50 mM NaF] in the presence of protease inhibitors.
Immunoprecipitation was performed by incubating the extracts with the M2-Flag
monoclonal antibody (Sigma) coupled to protein A Sepharose CL 4B at 4°C
for 1 hour. The precipitates were then washed with lysis buffer and subjected
to western blot analysis using an anti-HA antibody (Santa Cruz
Biotechnology).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
|
|
Convergent extension movements have been analyzed in a simple assay in
which naive AC cells can elongate in response to activin; this is associated
with the induction of cell populations that give rise to notochord and muscle
(Smith and Howard, 1992). To
analyze the effects of Xgly4 on activin-induced AC elongation, Xgly4 mRNA or
Xgly4Mo was injected with activin mRNA into the animal pole and the AC assay
was performed. Although tissue elongation occurred in ACs that had been
injected with activin mRNA injection, the elongation in response to activin
was strongly inhibited by co-injection of higher amounts of Xgly4 mRNA or
Xgly4Mo. In addition, Xgly4Mo-induced inhibition was rescued with a low dose
of wild-type Xgly4 mRNA without the native 5' UTR region
(Fig. 5A). For the analysis of
mesoderm induction by activin, the expression levels of both Xbra and Xwnt11
were measured using RT-PCR. The expression of these genes was not affected by
the co-injection of Xgly4 mRNA or Xgly4Mo
(Fig. 5B), which suggests that
Xgly4 controls cell movements without regulating the transcription of
mesodermal genes (mesoderm induction).
|
|
|
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
It has been shown genetically that mutations of the Drosophila
Dally and Dlp proteins, or the zebrafish Kny protein perturb normal patterning
or cell movements controlled by Wg or Wnt, respectively. It has been proposed
that the proteoglycans Dally, Dlp and Kny act as components of a receptor
complex that serves as a co-receptor for Fz. In addition to genetic evidence,
we have been able to show successfully that Xgly4 physically interacts with
the Wnt11 ligand (Fig. 9A) and
the Fz7 receptor (Fig. 9B),
which activate the non-canonical Wnt/PCP pathway. These results suggest that
Xgly4 may present Wnt ligands to Fz receptors, in the same way as the FGF
low-affinity HSPG receptor does with FGF high-affinity signalling receptors
(Pellegrini et al., 2000;
Schlessinger et al., 1995
). We
also found that Xgly4 physically interacts with Wnt8. The question arising
from these results is whether Xgly4 affects the Xwnt8 signaling pathway. In
fact, the overexpression of Xgly4 inhibits the Xwnt8-induced second axis.
However, because Otx2, the anterior marker gene, was expressed in
Xgly4Mo-injected embryos (data not shown), head formation, except eye
formation (Fig. 4B), was not
affected by reducing Xgly4 levels using Xgly4Mo; this lack of affect on head
formation is similar to that in the zebrafish kny mutant
(Topczewski et al., 2001
). The
expression of XmyoD gene, which is regulated by Xwnt8, was also
unperturbed (Fig. 4C), and we
also found that Xgly4 transcripts were not co-expressed with Xwnt8 in gastrula
(Christian et al., 1991
;
Ku and Melton, 1993
;
Moon et al., 1993
;
Tada and Smith, 2000
). In
addition, given that Xwnt8 is an endogenous ligand of Xgly4 and that Xgly4
acts as a co-receptor for Wnt8, depletion of Xgly4 by morpholino
oligonucleotides is expected to result in a loss of Wnt8 activity, and in
anteriorization of embryo, because Wnt8 is considered to be an inhibitor of
head formation. These results suggest that Xgly4 is not involved in the
regulation of the Xwnt8 canonical pathway in vivo during Xenopus
gastrulation, but that it inhibits Xwnt8 activity only when it is
overexpressed beyond the physiological level. Nevertheless, as several HSPGs
have been shown to act as modulators of the canonical Wg/Wnt pathway in
different model systems (Alexander et al.,
2000
; Baeg et al.,
2001
; Tsuda et al.,
1999
), we cannot rule out the possibility that Xgly4 could
potentially act as a modulator of the canonical Wnt/ß-catenin pathway if
it is co-expressed with the corresponding ligands.
Although embryos in which Xgly4 protein levels were reduced exhibited
defects of the dorsal convergent movements, embryos with overexpressed Xgly4
also exhibited these defects. We found that overexpression of Xgly4 inhibited
the non-canonical Wnt/PCP pathway in the same way as the kny gene
product does (Topczewski et al.,
2001). In the Drosophila wing disc, glypicans can
regulate the Wg protein distribution in the extracellular spaces and, in some
contexts at least, block Wg signaling (Baeg
et al., 2001
). Taken together, these findings suggest that high
levels of glypicans may serve to prevent Wg/Wnt ligands from interacting with
the signaling receptor. This is supported by our finding that Xgly4 forms a
complex with Fz7 (Fig. 9A,B).
It is also known that cell-cell adhesion, by the interaction of integrin and
fibronectin, disturbs Dsh localization and then convergent extension movements
(Marsden and DeSimone, 2001
).
Because Xgly4 is an extracellular matrix component, Xgly4 may play a role in
cell-cell interactions. In addition, we also speculate that glypicans may
affect the non-canonical Dsh/Wnt signaling pathway by regulating the
distribution of Wnt ligands in extracellular spaces during gastrulation in
vertebrates.
A growing body of evidence indicates that HSPGs also have crucial roles in
the regulation of other growth factors, including FGF, TGFß and the BMPs
(Bernfield et al., 1999;
Tumova et al., 2000
). It is
interesting to note that Drosophila dally interacts genetically with
dpp, a fly ortholog of vertebrate BMP, in the formation of the
genital organ. Thus, we suspected that Xgly4 might regulate BMP function as
well as the function of Wnt5A/Wnt11 during Xenopus development.
However, we found no indication that gain- or loss-of-function of Xgly4
affected the DV patterning regulated by BMP, which is consistent with our
observation that Xgly4 does not affect Bmp4-induced gene expression in whole
embryos (Xvent-1 expression pattern; Fig.
4C) and in the AC (data not shown). This observation is supported
by an independent observation by Topczewski et al., from cell tracing
experiments, that the fates of lateral cells in kny mutants are
comparable to the fates of the cells in wild-type embryos
(Topczewski et al., 2001
). In
addition to the BMP ligands, we investigated the functional interactions of
Xgly4 with eFGF and Xenopus nodal-related ligand 1, which are known
to be expressed in Xenopus gastrulae. Because neither reducing the
levels of nor overexpressing Xgly4 affected BMP, FGF or nodal signals (data
not shown), it appears that impaired gastrulation movements are not a
consequence of the regulation of BMP, FGF or nodal signals. We speculate that
the functional interaction of Xgly4 with Wnt and other ligands depends on the
extracellular environment, which is altered in a temporally and spatially
controlled manner.
In Xenopus gastrulae, Xwnt5A, which can also activate the
Dsh-independent Wnt/Ca2+ signaling pathway, physically binds to
Xgly4 protein (Fig. 9A) and is
co-expressed with Xgly4 in the dorsal ectoderm
(Moon et al., 1993). In
zebrafish, the phenotype of kny is similar to slb and also
to pipetail, which represents a mutation of zebrafish Wnt5A
(Rauch et al., 1997
). The
mediators of the Ca2+-dependent Wnt signaling cascade act in
convergent extension movements (Kuhl et
al., 2001
; Wallingford et al.,
2001
) and in tissue separation
(Winklbauer et al., 2001
).
Given that Xgly4 is the Xenopus ortholog of kny,
one might expect that Xgly4 would affect the Ca2+-dependent Wnt
signaling pathway. However, this may be unlikely because, in the AC assay, the
inhibition of activin-induced elongation by Xgly4Mo was efficiently rescued by
Xdsh alone. This suggests that, at least in the AC, Xgly4 affects only the
non-canonical Wnt/PCP pathway.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Adler, P. N. (1992). The genetic control of tissue polarity in Drosophila. BioEssays 14,735 -741.[Medline]
Alexander, C. M., Reichsman, F., Hinkes, M. T., Lincecum, J., Becker, K. A., Cumberledge, S. and Bernfield, M. (2000). Syndecan-1 is required for Wnt-1-induced mammary tumorigenesis in mice. Nat. Genet. 25,329 -332.[CrossRef][Medline]
Asashima, M., Nakano, H., Shimada, K., Kinoshita, K., Ishii, K., Shibai, H. and Ueno, N. (1990). Mesoderm induction in early amphibian embryos by activin A (erythroid differentiation factor). Roux's Arch. Dev. Biol. 198,330 -335.
Axelrod, J. D. (2001). Unipolar membrane
association of Dishevelled mediates Frizzled planar cell polarity signaling.
Genes Dev. 15,1182
-1187.
Axelrod, J. D., Miller, J. R., Shulman, J. M., Moon, R. T. and
Perrimon, N. (1998). Differential recruitment of
Dishevelled provides signaling specificity in the planar cell polarity and
Wingless signaling pathways. Genes Dev.
12,2610
-2622.
Baeg, G. H., Lin, X., Khare, N., Baumgartner, S. and Perrimon,
N. (2001). Heparan sulfate proteoglycans are critical for the
organization of the extracellular distribution of Wingless.
Development 128,87
-94.
Bernfield, M., Gotte, M., Park, P. W., Reizes, O., Fitzgerald, M. L., Lincecum, J. and Zako, M. (1999). Functions of cell surface heparan sulfate proteoglycans. Annu. Rev. Biochem. 68,729 -737.[CrossRef][Medline]
Blumberg, B., Wright, C. V., De Robertis, E. M. and Cho, K. W. (1991). Organizer-specific homeobox genes in Xenopus laevis embryos. Science 253,194 -196.[Medline]
Boutros, M. and Mlodzik, M. (1999). Dishevelled: at the crossroads of divergent intracellular signaling pathways. Mech. Dev. 83,27 -37.[CrossRef][Medline]
Boutros, M., Paricio, N., Strutt, D. I. and Mlodzik, M. (1998). Dishevelled activates JNK and discriminates between JNK pathways in planar polarity and wingless signaling. Cell 94,109 -118.[Medline]
Brickman, M. C. and Gerhart, J. C. (1994). Heparitinase inhibition of mesoderm induction and gastrulation in Xenopus laevis embryos. Dev. Biol. 164,484 -501.[CrossRef][Medline]
Cho, K. W., Blumberg, B., Steinbeisser, H. and De Robertis, E. M. (1991). Molecular nature of Spemann's organizer: the role of the Xenopus homeobox gene goosecoid. Cell 67,1111 -1120.[Medline]
Christian, J. L., McMahon, J. A., McMahon, A. P. and Moon, R. T. (1991). Xwnt-8, a Xenopus Wnt-1/int-1-related gene responsive to mesoderm-inducing growth factors, may play a role in ventral mesodermal patterning during embryogenesis. Development 111,1045 -1055.[Abstract]
Darken, R. S., Scola, A. M., Rakeman, A. S., Das, G., Mlodzik,
M. and Wilson, P. A. (2002). The planar polarity gene
strabismus regulates convergent extension movements in Xenopus.
EMBO J. 21,976
-985.
Djiane, A., Riou, J., Umbhauer, M., Boucaut, J. and Shi, D.
(2000). Role of frizzled 7 in the regulation of convergent
extension movements during gastrulation in Xenopus laevis.Development 127,3091
-3100.
Harland, R. M. (1991). In situ hybridization: an improved whole-mount method for Xenopus embryos. Methods Cell Biol. 36,685 -695.[Medline]
Heisenberg, C. P., Tada, M., Rauch, G. J., Saude, L., Concha, M. L., Geisler, R., Stemple, D. L., Smith, J. C. and Wilson, S. W. (2000). Silberblick/Wnt11 mediates convergent extension movements during zebrafish gastrulation. Nature 405, 76-81.[CrossRef][Medline]
Hoppler, S., Brown, J. D. and Moon, R. T. (1996). Expression of a dominant-negative Wnt blocks induction of MyoD in Xenopus embryos. Genes Dev. 10,2805 -2817.[Abstract]
Hopwood, N. D., Pluck, A. and Gurdon, J. B. (1989). MyoD expression in the forming somites is an early response to mesoderm induction in Xenopus embryos. EMBO J. 8,3409 -3417.[Abstract]
Hotta, K., Takahashi, H. and Satoh, N. (1998). Expression of an ascidian gene in the tip of the tail of tail-bud-stage embryos. Dev. Genes Evol. 208,164 -167.[CrossRef][Medline]
Iemura, S., Yamamoto, T. S., Takagi, C., Uchiyama, H., Natsume,
T., Shimasaki, S., Sugino, H. and Ueno, N. (1998).
Direct binding of follistatin to a complex of bone-morphogenetic protein and
its receptor inhibits ventral and epidermal cell fates in early Xenopus
embryo. Proc. Natl. Acad. Sci. USA
95,9337
-9342.
Itoh, K. and Sokol, S. Y. (1994). Heparan
sulfate proteoglycans are required for mesoderm formation in Xenopus
embryos. Development
120,2703
-2711.
Jones, C. M., Kuehn, M. R., Hogan, B. L., Smith, J. C. and
Wright, C. V. (1995). Nodal-related signals induce axial
mesoderm and dorsalize mesoderm during gastrulation.
Development 121,3651
-3662.
Keller, R. (1991). Early embryonic development of Xenopus laevis. Methods Cell Biol. 36, 61-113.[Medline]
Keller, R., Shin, J. and Domingo, C. (1992). The patterning and functioning of protrusive activity during convergence and extension of the Xenopus organiser. Development Suppl. 81-91.
Kelly, G. M., Erezyilmaz, D. F. and Moon, R. T. (1995). Induction of a secondary embryonic axis in zebrafish occurs following the overexpression of beta-catenin. Mech. Dev. 53,261 -273.[CrossRef][Medline]
Ku, M. and Melton, D. A. (1993). Xwnt-11: a
maternally expressed Xenopus wnt gene.
Development 119,1161
-1173.
Kuhl, M., Geis, K., Sheldahl, L. C., Pukrop, T., Moon, R. T. and Wedlich, D. (2001). Antagonistic regulation of convergent extension movements in Xenopus by Wnt/beta-catenin and Wnt/Ca2+ signaling. Mech. Dev. 106, 61-76.[CrossRef][Medline]
Larabell, C. A., Torres, M., Rowning, B. A., Yost, C., Miller,
J. R., Wu, M., Kimelman, D. and Moon, R. T. (1997).
Establishment of the dorso-ventral axis in Xenopus embryos is presaged by
early asymmetries in beta-catenin that are modulated by the Wnt signaling
pathway. J. Cell Biol.
136,1123
-1136.
Lin, X. and Perrimon, N. (1999). Dally cooperates with Drosophila Frizzled 2 to transduce Wingless signalling. Nature 400,281 -284.[CrossRef][Medline]
Marsden, M. and DeSimone, D. W. (2001). Regulation of cell polarity, radial intercalation and epiboly in Xenopus: novel roles for integrin and fibronectin. Development 128,3635 -3547.[Medline]
Mlodzik, M. (1999). Planar polarity in the
Drosophila eye: a multifaceted view of signaling specificity and cross-talk.
EMBO J. 18,6873
-6879.
Moon, R. T., Campbell, R. M., Christian, J. L., McGrew, L. L.,
Shih, J. and Fraser, S. (1993). Xwnt-5A: a maternal Wnt that
affects morphogenetic movements after overexpression in embryos of Xenopus
laevis. Development 119,97
-111.
Nieuwkoop, P. D. and Faber, J. (1967).A Normal Table of Xenopus laevis . Amsterdam: North Holland.
Nishimatsu, S. and Thomsen, G. H. (1998). Ventral mesoderm induction and patterning by bone morphogenetic protein heterodimers in Xenopus embryos. Mech. Dev. 74, 75-88.[CrossRef][Medline]
Pellegrini, L., Burke, D. F., von Delft, F., Mulloy, B. and Blundell, T. L. (2000). Crystal structure of fibroblast growth factor receptor ectodomain bound to ligand and heparin. Nature 407,1029 -1034.[CrossRef][Medline]
Rauch, G. J., Hammerschmidt, M., Blader, P., Schauerte, H. E., Strahle, U., Ingham, P. W., McMahon, A. P. and Haffter, P. (1997). Wnt5 is required for tail formation in the zebrafish embryo. Cold Spring Harbor Symp. Quant. Biol. 62,227 -234.[Medline]
Schlessinger, J., Lax, I. and Lemmon, M. (1995). Regulation of growth factor activation by proteoglycans: what is the role of the low affinity receptors? Cell 83,357 -360.[Medline]
Shih, J. and Keller, R. (1992). Cell motility
driving mediolateral intercalation in explants of Xenopus laevis.Development 116,901
-914.
Shulman, J. M., Perrimon, N. and Axelrod, J. D. (1998). Frizzled signaling and the developmental control of cell polarity. Trends Genet. 14,452 -458.[CrossRef][Medline]
Smith, J. C. and Howard, J. E. (1992). Mesoderm-inducing factors and the control of gastrulation. Development Suppl.127 -136.
Smith, J. C., Price, B. M., Green, J. B., Weigel, D. and Herrmann, B. G. (1991). Expression of a Xenopus homolog of Brachyury (T) is an immediate-early response to mesoderm induction. Cell 67,79 -87.[Medline]
Sokol, S. Y. (1996). Analysis of Dishevelled signalling pathways during Xenopus development. Curr. Biol. 6,1456 -1467.[Medline]
Suzuki, A., Kaneko, E., Maeda, J. and Ueno, N. (1997). Mesoderm induction by BMP-4 and -7 heterodimers. Biochem. Biophys. Res. Commun. 232,153 -156.[CrossRef][Medline]
Tada, M. and Smith, J. C. (2000). Xwnt11 is a
target of Xenopus Brachyury: regulation of gastrulation movements via
Dishevelled, but not through the canonical Wnt pathway.
Development 127,2227
-2238.
Thomsen, G., Woolf, T., Whitman, M., Sokol, S., Vaughan, J., Vale, W. and Melton, D. A. (1990). Activins are expressed early in Xenopus embryogenesis and can induce axial mesoderm and anterior structures. Cell 63,485 -493.[Medline]
Topczewski, J., Sepich, S. D., Myers, C. D., Walker, C., Amores, A., Lele, Z., Hammerschmidt, M., Postlethwait, J. and Solnica-Krezel, L. (2001). The Zebrafish Glypican Knypek controls cell polarity during gastrulation movements of convergent extension. Dev. Cell 1,251 -264.[Medline]
Tree, D. R., Shulman, J. M., Rousset, R., Scott, M. P., Gubb, D. and Axelrod, J. D. (2002). Prickle mediates feedback amplification to generate asymmetric planar cell polarity signaling. Cell 109,371 -381.[Medline]
Tsuda, M., Kamimura, K., Nakato, H., Archer, M., Staatz, W., Fox, B., Humphrey, M., Olson, S., Futch, T., Kaluza, V. et al. (1999). The cell-surface proteoglycan Dally regulates Wingless signalling in Drosophila. Nature 400,276 -280.[CrossRef][Medline]
Tumova, S., Woods, A. and Couchman, J. R. (2000). Heparan sulfate proteoglycans on the cell surface: versatile coordinators of cellular functions. Int. J. Biochem. Cell Biol. 32,269 -288.[CrossRef][Medline]
von Dassow, G., Schmidt, J. E. and Kimelman, D. (1993). Induction of the Xenopus organizer: expression and regulation of Xnot, a novel FGF and activin-regulated homeo box gene. Genes Dev. 7,355 -366.[Abstract]
Wallingford, J. B. and Harland, R. M. (2001).
Xenopus Dishevelled signaling regulates both neural and mesodermal convergent
extension: parallel forces elongating the body axis.
Development 128,2581
-2592.
Wallingford, J. B., Rowning, B. A., Vogeli, K. M., Rothbacher, U., Fraser, S. E. and Harland, R. M. (2000). Dishevelled controls cell polarity during Xenopus gastrulation. Nature 405,81 -85.[CrossRef][Medline]
Wallingford, J. B., Ewald, A. J., Harland, R. M. and Fraser, S. E. (2001). Calcium signaling during convergent extension in Xenopus. Curr. Biol. 11,652 -661.[CrossRef][Medline]
Wilson, P. and Keller, R. (1991). Cell rearrangement during gastrulation of Xenopus: direct observation of cultured explants. Development 112,289 -300.[Abstract]
Winklbauer, R., Medina, A., Swain, R. K. and Steinbeisser, H. (2001). Frizzled-7 signalling controls tissue separation during Xenopus gastrulation. Nature 413,856 -860.[CrossRef][Medline]
Yamamoto, T. S., Takagi, C. and Ueno, N. (2000). Requirement of Xmsx-1 in the BMP-triggered ventralization of Xenopus embryos. Mech. Dev. 91,131 -141.[CrossRef][Medline]
Yamanaka, H., Moriguchi, T., Masuyama, N., Kusakabe, M.,
Hanafusa, H., Takada, R., Takada, S. and Nishida, E.
(2002). JNK functions in the non-canonical Wnt pathway to
regulate convergent extension movements in vertebrates. EMBO
Rep. 3,69
-75.