1 Organogenesis and Neurogenesis Group, Center for Developmental Biology, RIKEN,
Kobe 650-0047, Japan
2 Department of Medical Embryology, Graduate School of Medicine, Kyoto
University, Kyoto 606-8501, Japan
* Author for correspondence (e-mail: sasaicdb{at}mub.biglobe.ne.jp)
Accepted 10 March 2005
![]() |
SUMMARY |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: Neural Crest, Pax3, Zic1, Wnt, Fate determination
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
A number of regulatory molecules in neural crest development have been
identified and appear to participate in the `multiple-step' fate determination
of this lineage (Aybar and Mayor,
2002; Knecht and
Bronner-Fraser, 2002
; Villianueva et al., 2002;
Meulemans and Bronner-Fraser,
2004
). However, the mechanism of how the initial determination of
the neural crest fate occurs at the restricted area of the ectoderm remains to
be understood. It has been suggested that graded BMP signals along the
dorsal-ventral (DV) axis play a role in the DV specification of the ectoderm
(the neural plate, the neural crest and the epidermis)
(Sasai and De Robertis, 1997
;
Dale and Jones, 1999
;
Mayor and Aybar, 2001
). Wnt
factors, which are expressed in the dorsal neural tube, are also implicated in
the promotion of neural crest development
(Wolda et al., 1993
;
Saint-Jeannet et al., 1997
;
Garcia-Castro et al., 2002
).
Nevertheless, it is poorly understood how these factors define the exact
boundaries of the neural plate, the neural crest and the epidermis in the
dorsolateral ectoderm. In addition, it is unclear whether these factors
promote the initial determination of the neural crest or the
maintenance/consolidation of differentiation.
Along the anteroposterior (AP) axis, the anterior limit of the neural crest corresponds to the anterior midbrain level. The forebrain level is devoid of typical neural crest tissues and is surrounded by specialized ectodermal tissues, such as the preplacode regions. In Xenopus, the cephalic neural crest anlage appears relatively early during embryogenesis, and neural crest-specific markers are detected even at the mid-gastrula stage. Moreover, in the frog research, neural crest determination can also be analyzed by using isolated ectodermal explants (animal cap assay), demonstrating that Xenopus is a suitable system to study the early phase of ectodermal specification into the neural crest fate.
The transcription factors Foxd3 and Slug are early bona fide markers of the
presumptive neural crest region in Xenopus, and play essential roles
in the specification of the neural crest fate in frog
(LaBonne and Bronner-Fraser,
1998; LaBonne and
Bronner-Fraser, 2000
; Sasai et
al., 2001
). In this study, we have investigated upstream
regulations of neural crest differentiation, particularly by focusing on the
roles of the transcription factors Pax3
(Bang et al., 1999
) and Zic1
(Kuo et al., 1998
;
Mizuseki et al., 1998
;
Nakata et al., 1998
). Mouse
genetic studies have indicated that Pax3
(Goulding et al., 1991
) is an
essential regulator of neural crest development
(Gruss and Walther, 1992
). The
Pax3 mutant (splotch) mouse exhibits defects in neural crest
derivatives, such as pigment cells, peripheral ganglia and cardiac neural
crest-derived structures (Epstein et al.,
1991
; Tassabehji et al.,
1992
; Conway et al.,
1997
). The human Waadernburg syndrome, which is caused by a
mutation in Pax3, also involves pigmentation defects
(Tassabehji et al., 1992
).
However, knowledge about the molecular function of Pax3 during early neural
crest development is still limited. In particular, the role of Pax3 in the
initial step of neural crest determination is largely unknown. Zic1 has been
implicated in the regulation of neural induction and neural crest development
(Kuo et al., 1998
;
Mizuseki et al., 1998
;
Nakata et al., 1998
). However,
because Zic1 is expressed in wider areas than the presumptive neural
crest (such as the anterior neural fold)
(Mizuseki et al., 1998
) (and
see below), the expression of Zic1 alone cannot explain the spatially
restricted pattern of neural crest development.
In this study, we examine the hypothesis that co-activation of Pax3 and Zic1 genes is the decisive event for the initiation of neural crest differentiation in the Xenopus ectoderm.
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
MO experiments
The morpholino antisense oligonucleotides (MOs) were designed as follows (a
lowercase letter indicates a mismatch):
In this study, `Pax3-MO' is the 1:1 mixture of Pax3A-MO and Pax3B-MO. The MOs do not contain sequences complimentary to the RNAs used in the rescue experiments. ß-catenin-MO was purchased from Gene Tools.
Embryonic manipulation and in situ hybridization
The developmental stage of the Xenopus embryos was determined
according to the normal table of Nieuwkoop and Faber
(Nieuwkoop and Faber, 1967).
For microinjection studies, synthetic mRNA (produced using mMESSAGE mMACHINE;
Ambion) and MOs were injected into two adjacent left animal blastomeres or two
ventral blastomeres of eight-cell-stage embryos. The injected embryos were
fixed with MEMFA (Sive et al.,
2000
) at the neurula stages. For animal cap assays, synthetic
mRNAs were injected into all animal blastomeres of eight-cell embryos, and
animal cap explants were prepared at stage 9 and cultured in 1 x Low
calcium and magnesium Ringer's solution (LCMR) with 0.1% BSA until stage 15.
Whole-mount in situ hybridization analyses were performed as previously
described (Sasai et al.,
2001
). For double in situ hybridization, signals with a
biotin-labeled probe were visualized by using BCIP red (magenta; BIOSYNTH AG)
and without NBT, and signals with a digoxigenin (DIG)-labeled probe were
visualized with BM purple (indigo; Roche).
Dissociated animal caps and RT-PCR analysis
For dissociated animal cap assays, animal cap explants were prepared from
injected embryos at stage 9, subsequently dissociated in the calcium- and
magnesium-free medium (Sive et al.,
2000) supplemented with 0.1% BSA, and cultured on four-well plates
(Nalge Nunc International) until stage 15. The dissociated animal cap cells
were treated with 100 ng/ml of recombinant mouse Wnt3a protein (R&D
Systems) during the indicated period. To exclude the possibility of
dissociation-induced artifacts, Foxd3 induction by Pax3
injection and Wnt3a protein treatment was also confirmed in undissociated
animal caps from which the impermeable outer layers were stripped. Total RNA
was extracted by using the RNeasy Micro kit (QIAGEN) and RT-PCR was performed
as previously described (Mizuseki et al.,
1998
; Sasai et al.,
2001
). The PCR primers used in this study for the first time were
nrp (forward 5'-TCACGACATGAGCTGGACTC-3', reverse
5'-CACAAACCCGAATCCTCTGT-3') and Pax3 3'UTR (forward
5'-TTTACCCGTTACTCATGGATAGTGT-3', reverse
5'-AATGTCACATAAAATCCAAAAAGGA-3'),
Western blot
Total proteins of injected or treated animal caps were extracted by
dissolving in extraction buffer [10 mM Tris-HCl (pH 7.4), 1% NP40, and
protease inhibitor cocktail (SIGMA)]. The cell extract (50 µg) was
subjected to SDS-polyacrylamide gel electrophoresis. Western blot analysis was
performed by using the anti-FLAG M2 antibody (SIGMA), the anti-HA antibody
(Roche), and ECL western blotting detection reagents (Amersham).
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
Next, we investigated possible signals that controlled the anterior limit
of Pax3 and Foxd3 expression. Because a previous study had
indicated that Wnt signaling plays a positive regulatory role in Pax3
expression (Bang et al., 1999),
we examined Wnt signaling by comparing its effects on the expression of
Pax3 and Foxd3. Unilateral injection of the
Wnt3a-expressing plasmid caused significant `rostral' expansion of
Pax3 and Foxd3 (67%, n=39 and 84%, n=38,
respectively; Fig. 1M,N),
whereas both genes were suppressed in embryos injected with dominant-negative
Tcf3 (dnTcf3, which blocks canonical Wnt signals)
(Molenaar et al., 1996
) mRNA
(Pax3, 77%, n=26; Foxd3, 67%, n=27; not
shown). These findings support the idea that Wnt signaling plays a crucial
role in the spatial regulation of the Pax3 and Foxd3
expression domains along the AP axis.
Collectively, expression of the early neural crest markers is closely associated with the overlapping expression of Pax3 and Zic1, both in normal embryos and in embryos with modified DV and AP patterns.
Overexpression of Pax3 and Zic1 induces neural crest differentiation in the embryonic ectoderm
To understand the causal relationship among Pax3, Zic1 and neural
crest differentiation, we next performed gain-of-function studies by mRNA
injection into two unilateral animal blastomeres of eight-cell embryos.
Overexpression of either Pax3 and Zic1 alone expanded
Foxd3 (65%, n=48 for Pax3; 74%, n=65 for
Zic1) and Slug (63%, n=51 for Pax3; 79%,
n=66 for Zic1) expression in the dorsolateral ectoderm
(Fig. 2A,B,D,E). In particular,
Pax3 injection frequently induced Foxd3 and Slug
expression in the anterior neural ridge region (48%, n=48 and 41%,
n=51, respectively; arrows in Fig.
2A,B), where neither the neural crest markers nor Pax3 is
normally expressed (note that Zic1 is normally expressed there, as
shown in Fig. 1B).
|
Misexpression of either Pax3 or Zic1 alone frequently caused ectopic expansion of Foxd3 and Slug within the dorsolateral region of the ectoderm, as shown above, but rarely in the ventral region (<5%; not shown). Similarly, when injected separately, Pax3 and Zic1 moderately expanded the expression of each other in the dorsolateral region (51%, n=55 and 66%, n=64, respectively; Fig. 2C,F), but not in the ventral ectoderm.
These findings prompted us to test whether co-expression of Pax3
and Zic1 induced ectopic neural crest differentiation in the ventral
ectoderm by injecting RNAs into either the ventral animal blastomeres of the
eight-cell stage (Fig. 2M-P),
or the ventral-most animal blastomeres of 16-cell stage embryo (see Fig. S1H
in the supplementary material). Single injection of Pax3 did not
induce substantial induction of either Foxd3 or Zic1 in the
ventral ectoderm (Fig. 2O and
Fig. S1G; data not shown). Similarly, Zic1 injection alone did not
induce either Foxd3 or Pax3 on the ventral side
(Fig. 2P and Fig. S1H; data not
shown). When injected together, Pax3 and Zic1 caused ectopic
expression of Foxd3 and Slug ventrally at a distance from
their orthotopic sites of expression (71%, n=42 and 60%,
n=55, respectively; Fig.
2M,N, arrow, and Fig. S1H). In this condition, little ectopic
expression of the neural marker nrp1 was induced ventrally
(n=27), whereas the epidermal marker Keratin was suppressed
(52%, n=25; not shown). The combined mRNA injection did not induce
ectopic expression of the preplacodal marker Six1
(Brugmann et al., 2004) (not
shown), or the dorsal neural tube marker 308a
(Tsuda et al., 2002
) (not
shown).
These findings demonstrate that co-activation of Pax3 and Zic1 is sufficient to induce ectopic neural crest differentiation in vivo, even in the ventral ectoderm, at the cost of epidermal differentiation.
Both Pax3 and Zic1 are required for neural crest determination in the embryo
We next investigated the roles of Pax3 and Zic1 in normal
development of the neural crest by performing loss-of-function studies using
morpholino antisense oligonucleotides (MOs), which inhibit the translation of
Pax3 and Zic1, respectively (see Fig. S2 in the
supplementary material). Injection of Pax3-MO suppressed the
expression of Foxd3 and Slug (suppression in 75%,
n=60 and 72%, n=54, respectively;
Fig. 3A,B), whereas injection
of the five-base-mispaired control MO did not (n=28 and
n=29, respectively; not shown). Suppression of Foxd3 was
reversed by co-injecting wild-type Pax3 mRNA (no suppression in 64%,
n=35; Fig. 3C).
Interestingly, Zic1 expression was not suppressed in the
Pax3-MO-injected embryo (n=38;
Fig. 3D; rather it was
upregulated, probably because of some feed-back mechanisms), indicating that
loss of the Pax3 function inhibits neural crest differentiation even in the
presence of Zic1 expression. The suppression of Foxd3 by
Pax3-MO was not reversed by overexpression of Zic1
(n=42; Fig. 3E) or
Msx1 (n=23; Fig.
3F).
|
We next examined whether Pax3 and Zic1 were required for the upregulation of Foxd3 expression caused by attenuation of BMP signaling (see Fig. 1L). Injection of Pax3-MO and/or Zic1-MO reversed the expansion of Foxd3 expression caused by dnBMPR (Fig. 3S-U, and data not shown), suggesting that attenuated BMP signaling requires both Pax3 and Zic1 for its enhancing effect on neural crest differentiation in vivo.
Requirement of Pax3 and Zic1 for neural crest differentiation in the animal cap explant
To further understand the mechanism of Pax3 and Zic1 functions in neural
crest differentiation, we next studied ectodermal explants. Unlike the
observation of the in vivo study (Fig.
2A), overexpression of Pax3 alone caused little induction
of Foxd3 in the animal cap explants (n=37;
Fig. 4A), suggesting that
animal caps lack some signals that are necessary for Pax3 to induce
Foxd3. We therefore tested co-injection of Wnt3a, which has
been shown to promote neural crest differentiation
(LaBonne and Bronner-Fraser,
1998). Co-injection of Pax3 and Wnt3a (but not
Wnt3a alone, Fig. 4B
inset) induced strong expression of Foxd3 (67%, n=42;
Fig. 4B), and also of
Zic1 (75%, n=24; Fig.
4C), in the animal caps, whereas neither Six1 nor
308a were induced (data not shown). The strong Foxd3
induction was inhibited by co-injecting Zic1-MO (n=32,
Fig. 4D).
|
We next investigated whether the induction of neural crest differentiation
by Pax3 and Zic1 was affected by enhanced BMP signaling.
Foxd3 induction by Pax3 and Wnt3a was strongly
inhibited by co-expression of Bmp4 (4%, n=25;
Fig. 4B,C,I,J; the level of
Bmp4 was sufficient to suppress neural induction by Chordin;
Fig. 4O,P). Similarly, the
induction of Foxd3 and Pax3 by Zic1 and
Wnt3a was reversed by Bmp4 (expansion in 6%, n=15,
and 17%, n=18, respectively; Fig.
4F,G,K,L). By contrast, Foxd3 induction by the
combination of Pax3, Zic1 and Wnt3a (86%, n=28;
Fig. 4M) was not remarkably
affected by the presence of BMP4 signals (67%, n=27;
Fig. 4N). These findings
indicate that co-presence of Pax3 and Zic1 initiates neural
crest differentiation in Wnt3a-treated animal cap ectoderm regardless of BMP
signaling. These observations are consistent with the in vivo observation that
injection of both Pax3 and Zic1 (but not each alone) induced
Foxd3 on the ventral side, where BMP signals are high
(Fainsod et al., 1994).
Neural crest development involves multiple determination steps and is
influenced by complex tissue interactions, such as the one between the neural
plate and epidermis (Knecht and
Bronner-Fraser, 2002). Therefore, one question as to the mode of
action for Pax3 and Zic1 is whether they work together in the precursors of
the neural crest or cooperate in a non-cell-autonomous manner by functioning
in different kinds of cells. To understand the cell-autonomous nature of the
differentiation control, we examined Pax3-induced neural crest
differentiation by using dissociated animal cap cells. We excised the animal
caps at stage 9 and dissociated them into single cells in calcium- and
magnesium-free Ringer solution (Fig.
4Q). The dissociated cells were cultured in the presence of Wnt3a
protein (added at the time equivalent to embryonic stage 9-13; harvested when
siblings reached stage 15). Foxd3 expression was induced in the
dissociated Pax3-injected animal caps when the Wnt treatment started
at stage 9 and 12, but not at stage 13
(Fig. 4R, lanes 5-7).
Pax3 injection and Wnt3a treatment induced strong Zic1 expression in dissociated animal caps (Fig. 4R, lanes 5, 6). We then investigated whether Zic1 was required for neural crest differentiation induced by Pax3 and Wnt3a in the dissociated animal caps. Co-injection of Zic1-MO inhibited Foxd3 expression induced by Pax3 and Wnt3a (Fig. 4S, lane 4), showing that Zic1 is essential for Pax3 to induce neural crest differentiation in dissociated animal cap cells under these conditions. Conversely, induction of Foxd3 and Pax3 by Zic1 and Wnt3a was also observed in dissociated animal cap cells (data not shown). These findings with the `dissociated' animal cap cells, in which non-autonomous regulation is unlikely to occur, support the idea that these two genes act together in a cell-autonomous manner.
Roles of Pax3, Zic1 and Wnt signals in neural crest determination in vivo
Pax3 and Zic1 are essential for the neural crest specification of the
ectoderm, both in the embryo and in the animal cap. In gain-of-function
experiments, Pax3 and Zic1 require the co-presence of exogenous Wnt signals to
evoke neural crest induction in the animal cap explant
(Fig. 4), but not in the embryo
(Fig. 2). Therefore, we next
tested whether endogenous Wnt signals were essential for Pax3 and
Zic1 to initiate neural crest differentiation in vivo. As shown in
Fig. 2, misexpression of both
Pax3 and Zic1 induces ectopic formation of neural crest
cells in the ventral ectoderm (Fig.
5A,B; red, lacZ tracer). This induction was significantly
suppressed when Wnt signaling was blocked by co-injection of
ß-catenin-MO (no significant induction observed, see
Fig. 5C,D, n=32 and
n=22, respectively) or dnTCF3 mRNA (no significant induction
observed, see Fig. S3C,D in the supplementary material, n=63 and
n=56, respectively). This suppression was reversed by additional
co-injection of wild-type ß-catenin (ectopic Foxd3 and
Slug induction in 36%, n=28 and 33%, n=27,
respectively; Fig. 5E,F) or
TCF3 (ectopic Foxd3 and Slug induction in 36%,
n=28 and 33%, n=27, respectively; Fig. S3E,F in the
supplementary material) mRNA. These findings indicate that neural crest
induction by Pax3 and Zic1 in the embryo is also dependent
on Wnt signaling.
|
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
In future investigation, the molecular dissection of the regulatory regions of the Pax3 and Zic1 genes would be very intriguing with regard to the `read-out' of the BMP activity gradient, and should be an attractive topic for promoter analyses using the transgenic frog technique. In addition, whether the co-expression of Pax3 and Zic1 directly attenuates BMP intracellular signaling should be examined to further clarify the mechanism of interactions.
The present study indicates two related but distinct roles of Wnt signals
for the initiation of neural crest differentiation. Wnt signaling is likely to
play a role in the control of the anterior limit of Pax3 expression
at early inductive phase (Fig.
1M,N) (Bang et al.,
1999). In addition, Wnt signaling has a cooperative function with
Pax3 and Zic1 factors for Foxd3/Slug induction, and for
mutual induction between Pax3 and Zic1
(Fig. 4). The earlier Wnt
function may be relevant to the `posteriorizing signals' in the double
gradient model (Villanueva et al.,
2002
). The later cooperative effect of Wnt could be interpreted in
line with the `lateralizing signals' of the two-signal model
(LaBonne and Bronner-Fraser,
1998
), which are suggested to enhance and reinforce neural crest
differentiation in weakly neuralized ectoderm. However, the exact relationship
between these functions needs to be clarified in future investigation. Also,
which particular Wnt factors act at each regulatory step in Xenopus
neural crest differentiation [such as Wnt6 and Wnt8 suggested for chick and
zebrafish neural crest induction
(Garcia-Castro et al., 2002
;
Lewis et al., 2004
)] is an
important question to be studied in future.
Recently, the role of FGF8 in neural crest differentiation has been
suggested with regard to paraxial mesoderm-derived inductive signals
(Monsoro-Burq et al., 2003).
Our preliminary study has indicated that Fgf8 also induces
Pax3 and Zic1 in vivo, and in the animal cap (see Fig. S4 in
the supplementary material). Fgf8-induced Foxd3 expression
in Chd-treated animal caps requires both Pax3 and Zic1, whereas the induction
of Pax3 and Zic1 themselves are not affected by
Zic1-MO and Pax3-MO, respectively (see Fig. S4D-J). These
findings suggest that FGF8 is another inductive signal candidate for
Pax3 and Zic1 expression.
In careful comparison, the Foxd3-expressing area appears to be
slightly narrower than the Pax3+/Zic1+
region (which includes the lateral-most part of the neural plate;
Fig. 1D-F, data not shown). One
interpretation for this could be that Foxd3 expression is inhibited
by certain neural plate-specific factors on the medial side. Another
possibility is that Wnt signaling, which is required for Pax3 and Zic1 to
induce Foxd3, is finely regulated by unknown local mechanisms. In
addition, more precise spatial regulation may be controlled by the interaction
of Pax3 and Zic1 with other transcription factors implicated
in neural crest development (e.g. Msx, Sox, Dlx, Myc and Ap2
genes) (Gammill and Bronner-Fraser,
2003; Meulemans and
Bronner-Fraser, 2004
).
Regarding the possible interaction with Msx1
(Suzuki et al., 1997b;
Tribulo et al., 2003
),
Msx1-MO suppresses Foxd3 expression without inhibiting
Pax3 and Zic1 expression in the neural crest region
(Fig. 3P-R). Conversely, the
attenuation of the Pax3 and Zic1 functions with MOs does not inhibit
Msx1 expression (data not shown), suggesting that
Msx1-mediated BMP signaling does not function upstream of
Pax3 and Zic1, but rather acts in an independent manner at
certain steps of neural crest differentiation. Consistently, unlike
Pax3 and Zic1, Msx1 does not induce Foxd3
expression in the animal cap even in the presence of Wnt3a (data not
shown). The understanding of the exact pathway network connecting Msx1 and
Pax3/Zic1/Wnt in neural crest differentiation requires further careful
consideration (Monsoro-Burq et al.,
2005
).
Do other Zic family members also participate in the initial step of neural
crest differentiation? In Xenopus, at least three members (Zic2,
Zic3 and Zic5) are expressed in overlapping patterns with
Zic1 (Nakata et al.,
1998; Nakata et al.,
2000
). Although these family members show moderately high homology
to Zic1 (54-57% identity of amino acid residues), the present study using the
specific MO has shown that Zic1 is indispensable for Xenopus neural
crest development. Interestingly, Zic1-MO injection does not suppress
the expression of Zic2, Zic3 and Zic5 in the neural crest
regions (see Fig. S5 in the supplementary material; instead, some moderate
upregulation was seen as shown by arrow), suggesting that these family genes
are not simply downstream of Zic1. It remains to be determined in
future whether the neural crest phenotype of Zic1 knockdown reflects
the quantitative change of total Zic-related activity in the presumptive
neural crest, or the qualitative differences of the role of Zic1 from the
others. In mice, the gene disruption of mouse Zic2 (but not mouse
Zic1) causes defects in neural crest development
(Aruga et al., 1998
;
Nagai et al., 2000
). The exact
roles of the Zic family members may be unambiguously studied by using reverse
genetics analyses, such as compound mutant mice.
Finally, a biochemical analysis of the cooperative function of Pax3 and Zic1 would be an intriguing and challenging topic for future study. Do they bind directly and cooperatively to the regulatory regions of target genes such as Foxd3? In our preliminary experiments, we have so far failed to detect co-immunoprecipitation of Pax3 and Zic1 proteins from the lysate of 293 cells overexpressing the two genes. Target DNA-dependent interactions of Pax3 and Zic1 proteins remain to be investigated and, for detailed study, must await the identification of their responsive elements in the regulatory regions of the Foxd3 and Slug genes.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
Footnotes |
---|
Supplementary material for this article is available at http://dev.biologists.org/cgi/content/full/132/10/2355/DC1
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Anderson, D. J. (1997). Cellular and molecular biology of neural crest cell lineage determination. Trends Genet. 13,276 -280.[CrossRef][Medline]
Aruga, J., Minowa, O., Yaginuma, H., Kuno, J., Nagai, T., Noda,
T. and Mikoshiba, K. (1998). Mouse Zic1 is involved
in cerebellar development. J. Neurosci.
18,284
-293.
Aybar, M. J. and Mayor, R. (2002). Early induction of neural crest cells: lessons learned from frog, fish and chick. Curr. Opin. Genet. Dev. 12,452 -458.[CrossRef][Medline]
Bang, A. G., Papalopulu, N., Goulding, M. D. and Kintner, C. (1999). Expression of Pax-3 in the lateral neural plate is dependent on a Wnt-mediated signal from posterior nonaxial mesoderm. Dev. Biol. 212,366 -380.[CrossRef][Medline]
Brugmann, S. A., Pandur, P. D., Kenyon, K. L., Pignoni, F. and
Moody, S. A. (2004). Six1 promotes a placodal fate within the
lateral neurogenic ectoderm by functioning as both a transcriptional activator
and repressor. Development
131,5871
-5881.
Conway, S. J., Henderson, D. J. and Copp, A. J.
(1997). Pax3 is required for cardiac neural crest
migration in the mouse: evidence from the splotch
(Sp2H) mutant. Development
124,505
-514.
Dale, L. and Jones, C. M. (1999). BMP signaling in early Xenopus development. BioEssays 21,751 -760.[CrossRef][Medline]
Epstein, D. J., Vekemans, M. and Gros, P. (1991). splotch (Sp2H), a mutation affecting development of the mouse neural tube, shows a deletion within the paired homeodomain of Pax-3. Cell 67,767 -774.[CrossRef][Medline]
Fainsod, A., Steinbeisser, H. and De Robertis, E. M. (1994). On the function of BMP-4 in patterning the marginal zone of the Xenopus embryo. EMBO J. 13,5015 -5025.[Abstract]
Gammill, L. S. and Bronner-Fraser, M. (2003). Neural crest specification: migrating into genomics. Nat. Rev. Neurosci. 4,795 -805.[CrossRef][Medline]
Garcia-Castro, M. I., Marcelle, C. and Bronner-Fraser, M.
(2002). Ectodermal Wnt function as a neural crest inducer.
Science 297,848
-851.
Goulding, M. D., Chalepakis, G., Deutsch, U., Erselium, J. R. and Gruss, P. (1991). Pax-3, a novel murine DNA binding protein expressed during early neurogenesis. EMBO J. 10,1135 -1147.[Abstract]
Gruss, P. and Walther, C. (1992). Pax in development. Cell 69,719 -722.[CrossRef][Medline]
Kishi, M., Mizuseki, K., Sasai, N., Yamazaki, H., Shiota, K.,
Nakanishi, S. and Sasai, Y. (2000). Requirement of
Sox2-mediated signaling for differentiation of early Xenopus
neuroectoderm. Development
127,791
-800.
Knecht, A. K. and Bronner-Fraser, M. (2002). Induction of the neural crest: a multigene process. Nat. Rev. Genet. 3,453 -461.[CrossRef][Medline]
Kuo, J. S., Patel, M., Gamse, J., Merzdorf, C., Liu, X., Apekin,
V. and Sive, H. (1998). opl: a zinc finger protein that
regulates neural determination and patterning in Xenopus.Development 125,2867
-2882.
LaBonne, C. and Bronner-Fraser, M. (1998).
Neural crest induction in Xenopus: evidence for a two-signal model.
Development 125,2403
-2414.
LaBonne, C. and Bronner-Fraser, M. (2000). Snail-related transcriptional repressors are required in Xenopus for both the induction of the neural crest and its subsequent migration. Dev. Biol. 221,195 -205.[CrossRef][Medline]
Le Douarin, N. M. and Kalcheim, C. (1999). The Neural Crest. Cambridge: Cambridge University Press.
Lewis, J. L., Bonner, J., Modrell, M., Ragland, J. W., Moon, R.
T., Dorsky, R. I. and Raible, D. W. (2004). Reiterated Wnt
signaling during zebrafish neural crest development.
Development 131,1299
-1308.
Marchant, L., Linker, C., Ruiz, P., Guerrero, N. and Mayor, R. (1998). The inductive properties of mesoderm suggest that the neural crest cells are specified by a BMP gradient. Dev. Biol. 198,319 -329.[CrossRef][Medline]
Matsui, M., Mizuseki, K., Nakatani, J., Nakanishi, S. and Sasai,
Y. (2000). Xenopus Kielin: a dorsalizing factor
containing multiple chordin-type repeats secreted from the embryonic midline.
Proc. Natl. Acad. Sci. USA
97,5291
-5296.
Mayor, R. and Aybar, M. J. (2001). Induction and development of neural crest in Xenopus laevis. Cell Tissue Res. 305,203 -209.[CrossRef][Medline]
Meulemans, D. and Bronner-Fraser, M. (2004). Gene-regulatory interactions in neural crest evolution and development. Dev. Cell 7,291 -299.[CrossRef][Medline]
Mizuseki, K., Kishi, M., Matsui, M., Nakanishi, S. and Sasai,
Y. (1998). Xenopus Zic-related-1 and Sox-2, two
factors induced by chordin, have distinct activities in the initiation of
neural induction. Development
125,579
-587.
Molenaar, M., ven de Wetering, M., Oosterwegel, M., Peterson-Maduro, J., Godsave, S., Korinek, V., Roose, J., Destree, O. and Clevers, H. (1996). XTcf-3 transcription factor mediates ß-catenin-induced axis formation in Xenopus embryos. Cell 86,391 -399.[CrossRef][Medline]
Monsoro-Burq, A. H., Fletcher, R. B. and Harland, R. M.
(2003). Neural crest induction by paraxial mesoderm in
Xenopus embryos requires FGF signals.
Development 130,3111
-3124.
Monsoro-Burq, A. H., Wang, E. and Harland, R. (2005). Msx1 and Pax3 cooperate to mediate FGF8 and WNT signals during Xenopus neural crest induction. Dev. Cell 8,167 -178.[CrossRef][Medline]
Nagai, T., Aruga, J., Minowa, O., Sugimoto, T., Ohno, Y., Noda,
T. and Mikoshiba, K. (2000). Zic2 regulates the kinetics of
neurulation. Proc. Natl. Acad. Sci. USA
97,1618
-1623.
Nakata, K., Nagai, T., Aruga, J. and Mikoshiba, K. (1998). Xenopus Zic family and its role in neural and neural crest development. Mech. Dev. 75, 43-51.[CrossRef][Medline]
Nakata, K., Koyabu, Y., Aruga, J. and Mikoshiba, K. (2000). A novel member of the Xenopus Zic family, Zic5, mediates neural crest development. Mech. Dev. 99,83 -91.[CrossRef][Medline]
Nguyen, V. H., Schmid, B., Trout, J., Connors, S. A., Ekker, M. and Mullins, M. C. (1998). Ventral and lateral regions of the Zebrafish gastula, including the neural crest progenitors, are established by a bmp2b/swirl pathway of genes. Dev. Biol. 199,93 -110.[CrossRef][Medline]
Nieuwkoop, P. D. and Faber, J. (1967).Normal Table of Xenopus laevis (Daudin) . Amsterdam: North Holland Publishing Company.
Saint-Jeannet, J. P., He, X., Varmus, H. E. and David, I. B.
(1997). Regulation of dorsal fate in the neuraxis by Wnt-1 and
Wnt-3a. Proc. Natl. Acad. Sci. USA
94,13713
-13718.
Sasai, N., Mizuseki, K. and Sasai, Y. (2001).
Requirement of FoxD3-class signaling for neural crest determination
in Xenopus. Development
128,2525
-2536.
Sasai, Y. and De Robertis, E. M. (1997). Ectodermal patterning in vertebrate embryos. Dev. Biol. 182,5 -20.[CrossRef][Medline]
Schmid, B., Furthauer, M., Connors, S. A., Trout, J., Thisse,
B., Thisse, C. and Mullins, M. C. (2000). Equivalent genetic
roles for bmp7/snailhouse and bmp2b/swirl in dorsoventral
pattern formation. Development
127,957
-967.
Sive, H. L., Grainger, R. M. and Harland, R. M. (2000). Early Development of Xenopus laevis: A Laboratory Manual. New York: Cold Spring Harbor Laboratory Press.
Suzuki, A., Thies, R. S., Yamaji, N., Song, J. J., Wozney, J.
M., Murakami, K. and Ueno, N. (1994). A truncated bone
morphogenetic protein receptor affects dorsal-ventral patterning in the early
Xenopus embryo. Proc. Natl. Acad. Sci. USA
91,10255
-10259.
Suzuki, A., Kaneko, E., Ueno, N. and Hemmati-Brivanlou, A. (1997a). Regulation of epidermal induction by BMP2 and BMP4 signaling. Dev. Biol. 189,112 -122.[CrossRef][Medline]
Suzuki, A., Ueno, N. and Hemmati-Brivanlou, A.
(1997b). Xenopus msx1 mediates epidermal induction and
neural inhibition by BMP4. Development
124,3037
-3044.
Tassabehji, M., Read, A. P., Newton, V. E., Harris, R., Balling, R., Gruss, P. and Strachan, T. (1992). Waardenburg's syndrome patients have mutations in the human homologue of the Pax-3 paired box gene. Nature 355,635 -636.[CrossRef][Medline]
Tribulo, C., Ayber, M. J., Nguyen, V. H., Mullins, M. C. and
Mayor, R. (2003). Regulation of Msx genes by a Bmp gradient
is essential for neural crest specification.
Development 130,6441
-6452.
Tsuda, H., Sasai, N., Matsuo-Takasaki, M., Sakuragi, M., Murakami, Y. and Sasai, Y. (2002). Dorsalization of the neural tube by Xenopus Tiarin, a novel patterning factor secreted by the flanking nonneural head ectoderm. Neuron 33,515 -528.[CrossRef][Medline]
Villanueva, S., Glavic, A., Ruiz, P. and Mayor, R. (2002). Posteriorization by FGF, Wnt, and retinoic acid is required for neural crest induction. Dev. Biol. 241,289 -301.[CrossRef][Medline]
Wada, H. (2001). Origin and evolution of the neural crest: a hypothetical reconstruction of its evolutionary history. Dev. Growth Differ. 43,509 -520.[CrossRef][Medline]
Wolda, S. L., Moody, C. J. and Moon, R. T. (1993). Overlapping expression of Xwnt-3A and Xwnt-1 in neural tissue of Xenopus laevis embryos. Dev. Biol. 155, 46-57.[CrossRef][Medline]