1 Howard Hughes Medical Institute and Department of Biological Chemistry,
University of California, Los Angeles, CA 90095-1662, USA
2 Victor Goodhill Ear Center, Head and Neck Surgery Division, University of
California, Los Angeles, CA 90095-1794, USA
3 Department of Cell Biology, Duke University Medical Center, Durham, NC 272710,
USA
4 Samuel Lunenfeld Research Institute, Mount Sinai Hospital, University of
Toronto, Toronto, M5G 1X5, Canada
Author for correspondence (e-mail:
bachiller{at}hnsurg.medsch.ucla.edu)
Accepted 24 April 2003
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SUMMARY |
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Key words: Chordin, Bmp, Tbx1, Fgf8, DiGeorge, Pharyngeal endoderm, Ventralization, Neural crest, Patterning, Persistent truncus arteriosus, Mouse
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INTRODUCTION |
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We report the loss-of-function mutation in the murine Chrd gene.
At day 8.5 of gestation (E8.5), a small fraction of homozygous mutant embryos
displayed a ventralized phenotype in which the allantois was expanded and the
embryonic region reduced in size. At gastrula and neurula stages Chrd
is expressed in the endomesoderm of the midline and in Hensen's node. Loss of
homozygotes was observed also at late gestation, but some Chrd
mutants survived and died of cardio-respiratory failure at birth. These
mutants closely mimic the alterations recently described for Tbx1
homozygous mouse mutants (Jerome and
Papaioannou, 2001; Lindsay et
al., 2001
; Merscher et al.,
2001
; Vitelli et al.,
2002a
) and recapitulate the pharyngeal malformations
characteristic of DiGeorge/Velo-Cardio-Facial (DGS/VCFS) syndromes
(Ryan et al., 1997
) in humans.
At mid embryogenesis, mouse Chrd is expressed in the endoderm of the
pharynx.
DGS/VCFS is the most common chromosomal microdeletion syndrome in humans,
affecting 1 in 4000 live births (Wilson et
al., 1994). DGS/VCFS is predominantly associated with
haploinsufficient microdeletions in human chromosome 22q11. DiGeorge syndrome
was initially identified in cases of isolated T cell immunodeficiency
(DiGeorge, 1968
;
Gatti et al., 1972
;
Harington, 1828-1829
), but
currently the term covers a spectrum of head and neck malformations, including
hypoplasia of thymus and parathyroid and thyroid glands, cleft palate, facial
dysmorphism with low setting of the external ear, small jaw, deafness, and
cardiac defects. The congenital heart malformations arise from incomplete
septation of the outflow tract (a defect frequently associated with defective
migration of the neural crest into the developing heart), and constitute the
primary cause of death in affected individuals
(Ryan et al., 1997
).
The defects observed in individuals with DGS/VCFS involve organs and
structures originating from the pharyngeal endoderm or adjacent tissues during
embryogenesis. Similar DiGeorge-like pharyngeal malformations are seen in
acquired syndromes caused by retinoic acid, alcohol or other in utero
teratogens (Ammann et al.,
1982; Lammer et al.,
1985
; Oster et al.,
1983
). Chrd mutant neonates display phenotypes
characteristic of DiGeorge syndrome: lack of thymus and parathyroid glands,
lack of heart colonization by neural crest, defects in pharyngeal arches 2-6,
cleft palate and abnormal placement of the external ear. Two different
developmental mechanisms have been proposed as possible explanations for the
pathogenesis of DiGeorge syndrome: incomplete differentiation of the
pharyngeal pouches (Wendling et al.,
2000
) and inability of peripharyngeal neural crest cells to
migrate to their target organs (Kirby and
Bockman, 1984
). Both processes are affected in Chrd
homozygous mouse mutants.
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MATERIALS AND METHODS |
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In situ hybridization, histological and skeletal preparations
Whole-mount in situ hybridization was performed as described
(Bachiller et al., 2000)
(http://www.hhmi.ucla.edu/derobertis/index.html).
Newborn and 14.5-day-old mouse embryos were fixed in Bouin's solution,
dehydrated, cleared and embedded in paraffin wax. Serial sections (8 µm)
were stained according to the Mallory's Tetrachrome method or with
Eosin/Hematoxylin. For sections shown in
Fig. 8, embryos were fixed in
paraformaldehyde for 4 hours after in situ hybridization, dehydrated, embedded
in Ducupan (Fluka) and sectioned at 10 µm. Alcian Blue/Alizarin Red
skeletal staining was performed as described
(Belo et al., 1998
).
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RESULTS |
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Heterozygous Chrd mice were viable and fertile and were mated to
generate Chrd-/- embryos of various developmental stages.
At day E8.5, we observed the presence of resorption nodules in the uterus of
pregnant females and a small reduction in the expected number of
Chrd-/- embryos (50 recovered, 57 expected). Four
genotyped homozygous mutant embryos showed a clear reduction in the size of
the embryonic region, accompanied by an enlargement of the allantois with
respect to the rest of the embryo (Fig.
2A,A'). In histological sections, a considerable hypoplasia
of the neural plate (Fig.
2B,B'), absence of somites and notochord
(Fig. 2C,C'), and an
abundance of extra-embryonic mesodermal cells in the allantois
(Fig. 2D,D') were
observed. The rest of the mutants (46) were morphologically indistinguishable
from their heterozygous and wild-type littermates. The phenotype of the four
abnormal mutants was similar to, but less pronounced than, the ventralization
of the mesoderm observed in double homozygous Chrd;Nog
mutants (Bachiller et al.,
2000) in which, in addition, anterior truncations of the neural
plate were also present.
|
Perinatal lethality
Only 49% (95 out of 194) of the expected Chrd-/-
animals were recovered at birth, all showing the same fully penetrant
phenotype. Of these, the majority was stillborn, but a few attempted,
unsuccessfully, to inflate their lungs. Externally, homozygous mutant neonates
were slightly smaller than their wild-type littermates and showed cyanosis,
microcephaly and reduction of the external ear, which was set abnormally close
to the eye (Fig. 3A').
Histological examination (Fig.
3B'-C') revealed the lack of thymus (t, a derivative
of the third pharyngeal pouch) and secondary palate (p), and hypoplasia of the
internal ear (ie) in the mutants. The anterior lobe and pars intermedia of the
pituitary gland (pi), both derived from the dorsal oral ectoderm immediately
adjacent to the cephalic border of the anterior endoderm, were normal
(Fig. 3C,C'). This
defined the rostral limit of the phenotype in the oropharynx, with
malformations restricted to derivatives of the Chrd-expressing
endoderm. The thyroid gland, which forms in ventral pharyngeal endoderm at the
foramen caecum, did differentiate but was hypoplastic and of irregular shape
(th, Fig. 3C'). The
parathyroid glands, derivatives of pharyngeal pouches 3 and 4, were absent
(data not shown), an observation consistent with the neonatal hypocalcaemia
seen in individuals with DiGeorge syndrome
(DiGeorge, 1968). We conclude
that the phenotype of Chrd-/- stillborn mice recapitulates
most of the features described in such individuals.
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The bones affected by the Chrd mutation have very different
origins. The basioccipital is purely of somitic origin; parts of the
basisphenoid arise from endochondral ossification of cephalic mesenchyme; the
palatine originates from intramembranous ossification of neural crest-derived
mesenchyme; the otic capsules differentiate from a mix of paraxial mesoderm
and neural crest cells; and the hyoid is strictly neural crest derived
(Le Douarin and Kalcheim,
1999). Amid such diversity of lineages, the unifying principle of
the phenotype seems to be the location of malformed structures in the
proximity of the Chrd-expressing axial mesendoderm
(Fig. 1E). This interpretation
is consistent with the observed premature degeneration of the anterior
notochord in Chrd-/- animals, and with the requirement of
prechordal plate and mesendoderm derived signals for the development of the
skeleton of the head (Belo et al.,
1998
; Couly et al.,
2002
; David et al.,
2002
).
DiGeorge-like cardiovascular defects
The cyanosis observed at birth can be a sign of cardiac malfunction. To
investigate this further, dissections at different stages of embryonic
development were performed. At E14.5, the hearts of
Chrd-/- animals showed a single vessel, instead of the
normal two, in the cardiac outflow tract
(Fig. 5D,D',E,E').
This condition is known in humans as persistent truncus arteriosus and is an
important malformation in individuals with DiGeorge syndrome. The lack of
separation between the ascending aorta and the pulmonary trunk may increase
the working load of the right ventricle causing its hypertrophy, as well as
the vasodilatation, oedema and haemorrhage seen in E14.5 embryos
(Fig. 5C'). As in
DiGeorge syndrome, defects in the cardiovascular system extended beyond the
outflow tract and included the great vessels derived from the pharyngeal arch
arteries (Fig. 6). In newborn
Chrd mutants, the common carotid arteries directly joined the truncus
arteriosus, resulting in the absence of the brachiocephalic artery and part of
the aortic arch (Fig. 6A-C). The pulmonary arteries originated directly from the proximal truncus
arteriosus, resulting in the absence of a common pulmonary trunk
(Fig. 6A-C). In addition,
laterality defects were observed, with an abnormal right turning of the aorta
in 40% of the mutants (Fig. 6, compare 6E
with 6F). When dissections were performed from the posterior, it
could be seen that, depending of the laterality of the descending aorta, the
right or left subclavian arteries adopted an abnormal retrooesophageal
position (Fig. 6D-F). Similar
defects have been described in chick embryos with neural crest cells ablations
(Kirby et al., 1983), and in
mice carrying deletions in the DiGeorge congenic region
(Lindsay et al., 1999
;
Merscher et al., 2001
) or
mutations in Tbx1 and Fgf8
(Abu-Issa et al., 2002
;
Frank et al., 2002
;
Jerome and Papaioannou, 2001
;
Lindsay et al., 2001
;
Vitelli et al., 2002b
).
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Pharyngeal abnormalities
To determine the onset of the pharyngeal phenotype we dissected pregnant
females from heterozygous matings at different times post coitum. At E9.0, a
stage at which Chrd is expressed in the pharyngeal endoderm,
Chrd-/- embryos could be identified by an indentation in
the neck region (Fig.
7A', arrow). The otic vesicles of the mutants were reduced
to half their normal diameter (Fig.
7A', arrowheads) and the second (hyoid) pharyngeal arch was
missing. Pharyngeal arches three to six never formed in mutant embryos
(Fig. 7B' and data not
shown). The missing or malformed structures are either direct precursors or
play inductive roles during the development of many of the organs that are
defective at birth in Chrd-/- mice. As most of the
phenotypic abnormalities observed in newborn mutants have their embryological
origin in the pharyngeal endoderm and the peripharyngeal region, we analyzed
the expression of a number of genes known to have important developmental
roles in human hereditary disease.
|
Next, we performed in situ hybridizations with Sox10, a gene
mutated in individuals with Waardenburg syndrome type 4
(Pingault et al., 1998) that
is expressed in neural crest and Schwann cells. At E7.5, the expression of
Sox10 was the same in Chrd-/- embryos and in
their wild-type littermates (data not shown). At E9.5, Sox10
expression in the dorsal root ganglia (drg) of the trunk was normal
(Fig. 7B,B'), but the
distribution of glial cells expressing Sox10 revealed specific
defects in the organization of the peripheral nervous system in the neck and
head region of the mutants (Fig.
7B'). In particular, cranial sensory ganglia showed marked
abnormalities. The trigeminal (tr) and vestibulo-cochlear (vc) ganglia,
corresponding to the V and VIII cranial nerves, respectively, were located
closer together in Chrd-/- embryos than in wild-type
littermates. In addition, abnormal nerve projections connecting the two of
them were seen (Fig. 7B',
arrowhead). The geniculate (g), petrosal (p) and nodose (n) ganglia,
corresponding to cranial nerves VII, IX and X, were the most affected, showing
either an extreme reduction in size or complete absence. These three ganglia
originate from the epibranchial placodes, and are known to require inductive
signals from anterior endoderm for their proper development
(Begbie et al., 1999
). The lack
of epibranchial placode-derived ganglia indicates that the secreted protein
Chrd is required for the activity of the inductive signal released by
pharyngeal endoderm.
Pax9 is a transcription factor required for the development of the
pharyngeal endoderm and its derivatives in the mouse
(Peters and Balling, 1999;
Peters et al., 1998
). At E9.5,
expression of Pax9 in the pharyngeal endoderm of
Chrd-/- embryos was weaker than in their wild-type
littermates (pe, Fig.
7D,D'). Pax9 expression revealed that the size and
shape of the pharynx was altered in the Chrd mutants, with pharyngeal
pouches reduced to a single swelling in the anterior-most region
(Fig. 7E,E'). The
hypoplasia of the pharynx was confirmed by histological sections of E14.5
embryos, in which the anterior endoderm appeared as a thin tube outlining a
greatly diminished lumen (ph, Fig.
5F,F'). The reduction of pharyngeal endoderm has also been
observed in Xenopus Chrd knockdowns
(Oelgeschläger et al.,
2003
). Non-pharyngeal regions in which Pax9 mRNA is
normally expressed, such as the somitic sclerotomes (sc) and facial mesenchyme
(fm), did not show differences in the distribution or abundance of the
transcripts (Fig.
7F,F').
We conclude from these studies that alterations in Pax3, Sox10 and Pax9 expression are restricted to a very limited area of wider expression domains, suggesting that lack of the secreted protein Chrd specifically disrupts local regulatory pathways acting in the peripharyngeal region surrounding the Chrd-expressing endoderm.
Tbx1 and Fgf8 expression requires chordin
To study the interaction of Chrd with genes known to cause
DiGeorge or DiGeorge-like phenotypes in mice, we analyzed the expression of
Tbx1 and Fgf8 in Chrd mutant embryos. Tbx1
is a member of the T-box family of transcription factors
(Papaioannou and Silver,
1998). It maps within the DGS/VCFS 22q11 microdeletion in humans
and has recently been shown to cause DiGeorge-like phenotype upon inactivation
in mice (Jerome and Papaioannou,
2001
; Lindsay et al.,
2001
; Merscher et al.,
2001
; Vitelli et al.,
2002a
). Expression of Tbx1 was altered in
Chrd-/- embryos. In wild-type E7.5 animals, Tbx1
is expressed in the foregut (future pharyngeal endoderm) and head mesoderm
(Fig. 8A). At this stage,
mutant littermates showed a clear reduction in the levels of Tbx1
expression in the same areas (Fig.
8A'). The reduction in Tbx1 mRNA was equally clear
in the pharyngeal region of Chrd homozygous embryos at E8.0, E8.5 and
E9.0 (Fig.
8B',C',D'). Transverse histological sections
showed that at the cellular level the abundance of Tbx1 transcripts
was drastically reduced in endoderm, both in the pharynx and foregut up to the
level of the hepatic diverticulum (Fig.
8F-H') Diminution in the concentration of Tbx1 mRNA
was also evident in mesoderm, including head, splanchnic (arrowheads) and
somatic mesoderm (arrows) in the peripharyngeal region
(Fig.
8F',G',H'). In addition, Tbx1
expression at E9 in the mesodermal core of the first pharyngeal arch was
diffuse, extending to most of the arch, and Tbx1 transcripts were
absent from the otic vesicle (Fig.
8D-D').
Fgf8 is a secreted growth factor expressed in a variety of
tissues, including the pharyngeal endoderm and neighboring mesoderm
(Crossley and Martin, 1995;
MacArthur et al., 1995
).
During early development, Fgf8 is required for gastrulation
(Sun et al., 1999
) and the
establishment of the left/right axis of symmetry
(Meyers and Martin, 1999
). At
later stages of Fgf8 is required for limb
(Lewandoski et al., 2000
;
Moon and Capecchi, 2000
) and
craniofacial (Trumpp et al.,
1999
) development. Recent experiments have shown that mice with
reduced Fgf8 activity present a spectrum of cardiovascular and
pharyngeal defects that closely mimic DiGeorge syndrome
(Abu-Issa et al., 2002
;
Frank et al., 2002
). In
addition, Fgf8 expression is abolished in the pharyngeal endoderm of
Tbx1-/- mutants and both genes interact genetically during
the differentiation of the pharyngeal arch arteries
(Vitelli et al., 2002b
). At
E9, Fgf8 expression in Chrd mutants is normal in the
mid-hindbrain isthmus, frontonasal prominence and tail. However, in pharyngeal
endoderm, Fgf8 transcript levels are drastically reduced
(Fig. 8E'). The reduction
of Tbx1 and Fgf8 expression in Chrd-/-
embryos suggested that both genes act downstream of Chrd in the same
regulatory pathway. These experiments do not determine whether Chrd
is required for the maintenance or for the induction of Tbx1 and
Fgf8 in the pharynx and neighboring tissues.
To test whether Chrd can induce Tbx1 and Fgf8, we injected Chrd mRNA (50 pg) into the ventral region of Xenopus embryos at the four-cell stage. Ventral marginal zone (VMZ) explants were dissected at early gastrula, cultured until sibling embryos reached early neurula stage, and analyzed by RTPCR. Tbx1 and Fgf8 mRNAs were expressed at high levels in whole embryos and dorsal marginal zone (DMZ) explants at this stage, and at low levels in VMZ explants (Fig. 8I, lanes 1-3). Upon microinjection, Chrd mRNA increased the levels of Tbx1 and Fgf8 in VMZ (Fig. 8I, lane 4). In situ hybridization of microinjected Xenopus embryos confirmed that the Tbx1 transcripts induced by Chrd mRNA were located in pharyngeal endoderm (data not shown). We conclude that Chrd, a Bmp antagonist, can induce Tbx1 and Fgf8 expression in Xenopus embryos, and is required for full expression of these genes in the pharyngeal region of the mouse embryo.
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DISCUSSION |
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Chrd and early development
As Chrd functions by regulating the access of Bmps to their receptors, the
effects of Chrd inactivation must result, at least initially, from
altered Bmp signaling. During late gastrulation Bmp2, Bmp4, Bmp5, Bmp6 and
Bmp7 are co-expressed in posterior mesoderm. Embryos that are homozygous
mutant for either Bmp2 or Bmp4, or double homozygous mutant for Bmp5 and Bmp7,
show severe reduction or absence of the allantois
(Fujiwara et al., 2001;
Solloway and Robertson, 1999
;
Zhang and Bradley, 1996
). This
phenotype is the opposite of the one described here, in which expansion of the
allantois at the expense of the embryonic mesoderm was observed in
Chrd-/- embryos at early somite stages
(Fig. 2). This suggests that
the lack of Chrd leads to an increase in Bmp signaling and a subsequent shift
in the differentiation of the trunk mesoderm towards a more ventroposterior
fate.
Chrd is not the only Bmp antagonist expressed at these stages.
Other proteins with possible Bmp antagonist activity, such as noggin
(McMahon et al., 1998),
follistatin (Iemura et al.,
1998
) and bambi (Grotewold et
al., 2001
) could collaborate in opposing the ventralizing activity
of Bmps. The existence of these potentially redundant genes may explain the
low penetrance or absence of ventralization observed when Bmp antagonists are
individually inactivated. In the case of Chrd;Nog double
homozygous mutants, ventralized embryos with large allantois were also
observed at the neural fold stage, although with more severe phenotypes
(Bachiller et al., 2000
). The
existence of a gastrulation phenotype in Chrd-/- embryos
indicates that the early functional compensation provided by Nog or
other Bmp inhibitors is not completely penetrant in Chrd mutants.
Chrd is required for pharyngeal development
The first manifestations of the post-gastrulation Chrd phenotype occur at
E8.5-9, at the pharyngula stage. At this time in development Chrd is
expressed in dorsal foregut and notochord
(Fig. 1E), and various Bmps are
expressed in the surrounding head and neck region
(Dudley and Robertson, 1997).
Bmps are potent growth factors involved in embryonic induction, cellular
differentiation and apoptosis. Their signals regulate the expression of
variety of transcription factors, among them several members of the Pax and
Tbx families (Peters and Balling,
1999
; Rodriguez-Esteban et
al., 1999
; Yamada et al.,
2000
). As a consequence of Chrd deficiency, the expression of
Tbx1 is reduced in the pharyngeal endoderm of
Chrd-/- mutants (Fig.
8).
We propose that Tbx1 mediates the autocrine effect of Chrd on the
endoderm, subsequently affecting the formation of the thymus, parathyroid
glands and other pharyngeal endoderm derivatives that are defective in
Tbx1-/- (Jerome and
Papaioannou, 2001; Lindsay et
al., 2001
; Merscher et al.,
2001
) and Chrd-/- mice (this work). As
development progresses, the endodermal component of the DiGeorge phenotype may
be aggravated by the reduction of Pax9 and Fgf8 expression
in pharyngeal endoderm (Fig.
7E',F'), for Pax9 mouse mutants lack
derivatives of the pharyngeal pouches
(Peters et al., 1998
), and
mice with reduced Fgf8 activity present DiGeorge-like phenotypes
(Abu-Issa et al., 2002
;
Frank et al., 2002
). It has
been shown that Fgf8 expression is eliminated from the endoderm of
Tbx1-/- mutants
(Vitelli et al., 2002b
), and
therefore the decrease in Tbx1 levels observed in
Chrd-/- embryos could explain the phenotype observed.
However, our experiments do not exclude the possibility that Chrd may
also control Fgf8 and other endoderm expressed genes through a
parallel Tbx1-independent route. Additional experiments will also be
required to explore the existence of a possible regulatory loop linking the
maintenance of Chrd expression to Tbx1 activity. The
disruption of endoderm development would in turn impair signaling to nearby
ectoderm (Begbie et al., 1999
),
preventing the induction of the epibranchial placodes, which are missing in
Chrd-/- embryos (Fig.
7B').
Chrd and skeletal development
The striking similarities between the Chrd-/- and
Tbx1-/- phenotypes, and the reduction of Tbx1
expression in the head mesoderm of Chrd mutants, suggest that
Tbx1 may also be a mediator of the paracrine effects of Chrd on
peripharyngeal mesoderm. Although at first inspection most of the defects
observed in this area seem to involve derivatives of the neural crest, the
phenotypes in the base of the skull and rostral vertebral column suggest that
some of the structures affected are of head mesoderm or somitic origin, and
thus derived from Tbx1-expressing paraxial mesoderm. A further
indication that some of the defects observed in the peripharyngeal region of
the Chrd-/- animals originate in the mesoderm
independently of the neural crest, is provided by a comparison with the
phenotype of the endothelin A receptor (Ednra) mutation in mouse
(Clouthier et al., 1998). Ednra
activity is cell autonomous in the neural crest, and upon disruption causes
cardiac and head and neck defects reminiscent of DGS/VCFS, but does not
produce malformations of the axial skeleton as reported here for Chrd
mutant animals.
Pharyngeal endoderm patterns the neural crest
One of the salient characteristics of DiGeorge syndrome is the presence of
persistent truncus arteriosus in the outflow tract of the heart. This
phenotype is also seen in mice homozygous for Sp2H, a
mutation in the Pax3 gene that affects neural crest migration
(Conway et al., 1997). In
Chrd homozygous mutant embryos, Pax3 expression is normal in
the cranial neural crest, while the cells are still located within the neural
folds, but at later stages the migration of Pax-3-positive neural
crest cells is impaired in the mutants. As Chrd is not expressed in
neural crest, the abnormalities observed must be secondary to the lack of
Chrd expression in pharyngeal endoderm. In zebrafish, oneeye
pinhead (oep), Casanova (cas) and Van
Gogh (vgo) mutants cause defects in the endoderm that interfere
with the correct migration of neural crest cells
(David et al., 2002
;
Piotrowski and Nusslein-Volhard,
2000
). In the chick, transplantation of pharyngeal endoderm has
shown that this tissue instructs Hox-negative neural crest cells to
differentiate into particular elements of the head skeleton
(Couly et al., 2002
). The
overall structure of the region through which the neural crest must migrate is
disorganized in Chrd-/- mutants. In this respect it should
be noted that pharyngeal arches 2 to 6 fail to form in these Chrd
mutants, indicating that Chrd may be particularly important for the patterning
of Hox-positive neural crest.
Pharyngeal malformations and Bmp signaling
Chrd maps outside the DiGeorge microdeletion (to human chromosome
3q27) (Pappano et al., 1998)
(D.B. and E.M.D.R., unpublished). As DGS/VCFS is not linked in all individuals
to deletions in the 22q11 region, the finding that mutations in Chrd
can reproduce DiGeorge syndrome in mice offers a new candidate gene for
genetic testing in humans. Chrd activity is modulated post-translationally by
metalloproteinases (Piccolo et al.,
1997
) and by other Bmp-binding proteins such as twisted
gastrulation (Oelgeschlager et al.,
2000
) and noggin (Bachiller et
al., 2000
). Allelic variation in any of these genes, or in
components of the Bmp signal transduction pathway could also potentially lead
to DiGeorge phenotypes. Furthermore, the realization that multiple components
of the Chrd/Bmp pathway may participate in DGS/VCFS might help to explain the
considerable phenotypic variability observed in humans and mice with
rearrangements in 22q11 (Ryan et al.,
1997
; Taddei et al.,
2001
).
The Chrd/Bmp/Noggin signaling system is essential for the establishment of
the three major body axes during gastrulation
(Bachiller et al., 2000). As
shown here, Chrd is also required for patterning the head and neck region of
the vertebrate embryo at the pharyngula stage, a time of maximal complexity in
the regulatory interactions taking place in the embryo
(Raff, 1996
). Many congenital
malformations have their origin at this particular time in development, which
corresponds to the phylotypic stage of the vertebrates
(Slack et al., 1993
). Further
analysis of pharyngeal development will provide a conceptual framework for
understanding the role of Chrd/Bmp signaling in the pathogenesis of DiGeorge
syndrome and other developmental defects arising in the head and neck region
during vertebrate development.
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ACKNOWLEDGMENTS |
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Footnotes |
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