1 Department of Craniofacial Development and Orthodontics, Kings College London,
Floor 28, Guy's Tower, Guy's Hospital, London SE1 9RT, UK
2 MRC Human Genetics Unit, Western General Hospital, Crewe Rd, Edinburgh EH4
2XU, UK
3 Center for Animal Resources and Development (CARD), Kumamoto University,
Kumamoto, Japan
Author for correspondence (e-mail:
robert.hill{at}hgu.mrc.ac.uk)
Accepted 2 December 2003
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SUMMARY |
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During embryogenesis, Bapx1 is expressed in a discrete domain within the mandibular component of the first branchial arch and later in the primordia of middle ear-associated bones, the gonium and tympanic ring. Consistent with the expression pattern of Bapx1, mouse embryos deficient for Bapx1 lack a gonium and display hypoplasia of the anterior end of the tympanic ring. At E10.5, expression of Bapx1 partially overlaps that of Gsc and although Gsc is required for development of the entire tympanic ring, the role of Bapx1 is restricted to the specification of the gonium and the anterior tympanic ring. Thus, simple overlapping expression of these two genes appears to account for the patterning of the elements that compose the structural components of the middle ear and suggests that they act in concert.
In addition, Bapx1 is expressed both within and surrounding the incus and the malleus. Examination of the malleus shows that the width, but not the length, of this ossicle is decreased in the mutant mice. In non-mammalian jawed vertebrates, the bones homologous to the mammalian middle ear ossicles compose the proximal jaw bones that form the jaw articulation (primary jaw joint). In fish, Bapx1 is responsible for the formation of the joint between the quadrate and articular (homologues of the malleus and incus, respectively) enabling an evolutionary comparison of the role of a regulatory gene in the transition of the proximal jawbones to middle ear ossicles. Contrary to expectations, murine Bapx1 does not affect the articulation of the malleus and incus. We show that this change in role of Bapx1 following the transition to the mammalian ossicle configuration is not due to a change in expression pattern but results from an inability to regulate Gdf5 and Gdf6, two genes predicted to be essential in joint formation.
Key words: Bapx1, Middle ear, Tympanic ring, Gonium, goosecoid, Evolution, Mouse
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Introduction |
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Current conjecture on the molecular changes that accompany evolution is
centred on changes within regulatory proteins. Analysis of molecular changes
that are crucial for morphological transitions are feasible in vertebrates as
integral changes in morphology that characterise a class are well documented.
The homeobox-containing transcription factors have been implicated as key
elements in regulatory changes and the data increasingly supports a role for
both single homeobox genes and whole complex Hox clusters in the evolution of
animal morphology (e.g. Grenier and
Carroll, 2000; Grandien and
Sommer, 2001
). Bapx1 (Nkx3.2) is an
evolutionarily conserved homeobox-containing gene that belongs to the NK-2
family of transcription factors (Kim and
Nirenberg, 1989
; Tribioli et
al., 1997
; Tribioli and
Lufkin, 1997
) and is most closely related to the Drosophila
bagpipe (Nkx3) gene. In vertebrates, craniofacial expression of
the gene is detected (Newman et al.,
1997
), including a large region of the intermediate first arch
that encompasses the jaw joint region. Recent work carried out by Miller et
al. (Miller et al., 2003
)
reveals a role for the gene in regulating the patterning of the zebrafish
(D. rerio) jaw joint. In this organism, Bapx1 is initially
expressed in the mesenchyme of the mandibular arch primordia but later can be
detected in the cells within and surrounding the jaw joint. Downregulation of
the gene, using Bapx1-specific morpholinos, results in a
dose-dependent loss of the jaw joint, and is characterised by a fusion between
the constituent cartilages, the quadrate and articular.
Given the evolutionary relationship between the jaw bones of fish and the
middle ear bones of mammals, these observations raise the possibility that
Bapx1 may play a crucial role in regulating the development of the
murine middle ear. Consistent with this hypothesis, expression of the gene has
been reported at E10.5 in the mandibular portion of the first branchial arch,
and later in precursor of Meckel's cartilage
(Tribioli et al., 1997).
Previous analyses of mice carrying targeted mutations in the Bapx1
gene have reported a role for the gene in regulating the development of the
axial skeleton, spleen and the gastroduodenal region of the gut
(Lettice et al., 1999
;
Tribioli and Lufkin, 1999
;
Akazawa et al., 2000
). We show
here that Bapx1 plays a crucial role in regulating the development of
the structural elements of the murine middle ear, and provide evidence to
suggest that it does so in combination with Gsc, a gene with a
well-characterised role in tympanic ring development. Furthermore, we
demonstrate that the role of Bapx1 in development of the middle ear
ossicles is restricted to regulating the width of the malleus and that the
lack of the predicted malleal/incal fusions in Bapx1-/-
mice results from regulatory changes in genes involved in joint formation.
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Materials and methods |
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Probes and in situ hybridisation
DIG in situ hybridisation was performed on whole-mount embryonic day (E)
10.5-12.5 mouse embryos and on sections derived from paraformaldehyde-fixed
and paraffin-embedded tissue essentially as described by Wilkinson
(Wilkinson, 1992). Radioactive
35S in situ hybridisation procedures were carried out as described
by Tucker et al. (Tucker et al.,
1999
). Following skeletal staining and conventional imaging,
samples were embedded in 2% LMP agarose made up in 80% glycerol and analysed
by OPT essentially as described (Sharpe et
al., 2002
). Because the samples had been cleared previously in
KOH/glycerol, the methanol and BABB pre-scan treatments were omitted and the
scans were performed in a cuvette filled with 80% glycerol.
Skeletal staining of mouse and chick embryos
E14.5 to postnatal day 2 (P2) mouse embryos were stained for bone and
cartilage formation using the method described by Kessel and Gruss
(Kessel and Gruss, 1991). E7
to E14 chick embryos were decapitated and heads fixed in 4% PFA overnight.
Heads were then washed in PBS and stained in Alcian Blue 8GX overnight (5% of
a 0.1% Alcian Blue solution, 5% acetic acid in ethanol). Stained heads were
rehydrated through an ethanol series over several days and then cleared in 1%
KOH. For staining of cranial bones, alcian blue heads were stained in 1%
Alizarin Red in 0.5% KOH overnight. Heads were washed in 0.5% KOH to remove
excess stain. Once cleared, heads were taken up through a glycerol series for
storage.
Statistical analysis
Middle ears were dissected from E18.5 wild-type (n=10),
heterozygous (n=13) and homozygous (n=8) mice. The
dimensions of the malleus were measured and pairwise Student's
t-tests were carried out on the data generated.
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Results |
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Bapx1 expression correlates with tympanic ring condensation and development of the gonium
Bapx1 expression has been previously reported in the inferior face
of the mandibular portion of the first branchial arch at E10.5, and later, at
E12.5, in a domain associated with Meckel's cartilage
(Tribioli et al., 1997)
(Fig. 4A,B). Our observations
support these findings but also reveal expression of Bapx1 at E12.5
in the mesenchymal condensation that ultimately gives rise to the tympanic
ring (Mallo and Grindley,
1996
) (Fig. 4C,D).
Strong expression of Bapx1 is also observed in a small mesenchymal
condensation medial to the developing tympanic ring and adjacent to Meckel's
cartilage that we postulate to be the prospective gonium
(Fig. 4D).
|
Bapx1 and Gsc function independently to specify the tympanic ring
Mouse studies have revealed numerous genes that, when inactivated, affect
the development of the bones associated with the middle ear including
Fgf8 (Trumpp et al.,
1999), Prx1 (Martin
et al., 1995
) and Msx1
(Satokata and Maas, 1994
).
Expression of each of these genes is unaffected at E10.5 in
Bapx1-/- mice (data not shown). Of those genes that give
rise to phenotypes that resemble those observed in
Bapx1-/- mice, Gsc is the best characterised.
Gsc is expressed at E10.5 in the mandibular component of the first
branchial arch and later in the region of the developing auditory meatus and
malleus (Gaunt et al., 1993
).
Accordingly, inactivation of the gene results in a variety of defects
associated with the middle ear that include aplasia of the tympanic cavity and
the external auditory meatus and defects in the manubrium and processus brevis
of the malleus. Significantly, Gsc-/- mice lack a tympanic
ring (Rivera-Pérez et al.,
1995
; Yamada et al.,
1995
) or display only a rudimentary element
(Kuratani et al., 1999
), and
are reported to exhibit mild hypoplasia of the gonial bone
(Rivera-Pérez et al.,
1995
; Yamada et al.,
1995
). Given the phenotypic similarities of the Bapx1 and
Gsc knockout mice, the possibility that the two genes might function
together to specify tympanic ring development was investigated. At E10.5, both
genes are expressed in most, perhaps all, cells within well-defined domains
within the mandibular component of the first branchial arch
(Fig. 5A,B). Two-colour in situ
hybridisation analysis reveals that these domains overlap within a small area
in the caudoproximal region of the arch
(Fig. 5C,D). Later in
development at E15.5, Gsc is expressed strongly throughout the
condensing tympanic (Fig.
5E,G,J,L) and around the EAM up to the malleus
(Fig. 5L), coinciding with
Bapx1 expression in the caudal region of the EAM
(Fig. 5K) and in the anterior
tympanic ring (Fig. 5E-G).
Gsc expression is only weakly seen in the developing gonium, where
Bapx1 is strongly expressed (Fig.
5F,G).
|
Comparison of tympanic ring and gonium development in mouse with prearticular and jaw development in chick
The mammalian tympanic ring is thought to be homologous to the angular of
non-mammalian gnathostomes, while the gonium is homologous to the prearticular
(Gaupp, 1911;
Voit, 1924
). These two
membranous bones are closely associated with the cartilaginous articular,
which (according to Reichert's theory) is homologous to the mammalian malleus
(Reichert, 1837
). An
investigation of the development of these two membranous bones in species that
represent the archetypal configuration of facial bones following
(Fig. 6A-C), and preceding
(Fig. 6D-F), the transition to
the middle ear was undertaken.
|
|
Expression of Eya1, Gdf5 and Gdf6 in developing mouse middle ear
The lack of a phenotype between the malleus and incus of
Bapx1-/- mice, as predicted by the zebrafish work and the
conserved expression pattern, was unexpected. This might be due to
compensation by the other member of the bagpipe family, Nkx3.1.
Double knockouts for Bapx1 and Nkx3.1 show an enhanced
skeletal phenotype compared to single Bapx1 mutants, demonstrating
that the two genes do collaborate in some aspects of embryonic development
(Herbrand et al., 2002).
Nkx3.1 is expressed in epithelial structures of the face, such as the
tongue and teeth but not the middle ear
(Tanaka et al., 1999
).
Expression of Nkx3.1 is unaffected in facial structures in the
Bapx1 mutant (Fig. 8A)
and no upregulation is observed in the branchial arches or middle ear, either
at E10.5, E14.5 or E15.5 (Fig.
8B,C and data not shown). The failure to get a defect in formation
of the incudo-malleal joint in Bapx1-/- embryos cannot
therefore be attributed to compensation by Nkx3.1 in this region.
|
In zebrafish, the reduction of Bapx1 expression results in loss of
Gdf5 expression in the developing jaw joint. Gdf5 encodes a
TGFß-related signalling factor expressed in early cartilage condensations
and later in developing joints (Storm et
al., 1994). A subset of mouse appendicular and axial joints
requires GDF5 function (Storm and Kingsley, 1996). Therefore Gdf5
expression was examined in the developing joint region in between the malleus
and incus in the mouse. At E15.5, when these two cartilages are clearly
separated, Gdf5 expression was observed between the incus and
malleus, overlapping the expression of Bapx1
(Fig. 8G,I). Gdf5
expression was therefore examined in the Bapx1 mutant mice.
Surprisingly, given the zebrafish result, no effect on Gdf5
expression could be seen (Fig.
8J). Gdf6 was recently shown to be expressed between the
incus and malleus during middle ear development and Gdf6 mutant mice
have defects in the middle ear articulations not previously seen for
Gdf5 mutants (Settle et al.,
2003
). Analysis of the expression of these two Gdf genes in the
chick showed that both Gdf5 and Gdf6/7 (a single gene with
homology to both murine Gdf6 and Gdf7) were expressed in the
joint region between the quadrate and articular
(Wilson and Tucker, 2004
)
(data not shown). In the mouse, Gdf6 expression pattern was similar
to Gdf5 in the developing middle ear
(Fig. 8H). No change in
Gdf6 expression, however, was observed in the Bapx1 mutant
mice (Fig. 8K). Therefore, the
joint signalling cascade controlled by Bapx1, as identified in
zebrafish, does not appear to be active in the mammalian middle ear. Continued
expression of joint markers in between the malleus and incus can therefore
explain the lack of a defect in this region in the mutant mice.
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Discussion |
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Our findings raise the possibility of a developmental pathway regulated by
Bapx1 that is specific for defining the width of the malleus. The
idea of several different developmental mechanisms that are each crucial for
regulating distinct aspects of the development of the middle ear ossicles has
been proposed (Mallo, 1997).
Administration of RA to embryos at different stages of gestation suggests a
temporal order to the patterning of the malleus
(Mallo, 1997
), although
genetic studies, including those involving targeting of Gsc, Msx1 and
Prx1, suggest the existence of discrete regulatory pathways for
domains within the ossicle. The functional significance of
malleal-width-specific pathway in the mouse is not immediately apparent;
however, it is tempting to speculate that changes in the structure of the
middle ear ossicles may have a bearing on aspects of an organisms hearing
capabilities, as has been demonstrated with respect to tympanic-membrane area
and middle ear structure (Rosowski,
1992
).
Link between presence of a gonium and frequency of hearing
Postnatally, the gonium fuses with both the tympanic ring and the malleus
and is referred to as the process folii (anterior or gracilis), a component of
the malleus. Clearly, however, the gonium is an independently specified
component of the middle ear. Studies into the auditory role of the gonium are
scarce; however, consensus suggests that the gonium acts as a rigid link
connecting the malleus to the tympanic bone
(Fleischer, 1978). Analysis of
audiograms taken from a variety of species suggest a relationship between the
degree of fusion between the malleus and the tympanic bone, as mediated by the
gonium, and the range of frequencies that the animal can detect
(Fleischer, 1978
;
Rosowski, 1992
). Animals with
a complete gonium fusion hear at the higher end of the frequency spectrum.
Those in which the gonium is underdeveloped or lacking entirely (e.g. human
and rabbit), have a correspondingly weak link between the malleus and tympanic
bone and consequently, are suggested to have an enhanced ability to hear lower
frequencies of sound. It is tempting to speculate therefore, that although not
rendering the mice deaf, the gonium hypoplasia observed in Bapx1
heterozygotes may have an effect on the range of frequencies that these mice
can detect. Furthermore, we suggest that expression levels of Bapx1
may be important in different mammalian species in defining the auditory
frequency range of detection.
Bapx1 and Gsc operate in combination in the gonium and tympanic ring
In recent years, expression patterns for a number of genes in the first
branchial arch have been described that appear highly regionalised and
overlapping, raising speculation that the specific combination of genes
expressed within any given region ultimately determines the developmental
fate. The expression of Bapx1 and Gsc is an example; these
genes are expressed from E10.5 in an overlapping pattern, Bapx1 being
more proximal than Gsc. Inactivation of either Bapx1 or
Gsc results in defects associated with the tympanic ring. Although
loss of Gsc results in aplasia of this skeletal element, only the
anterior-most thickening fails to develop in Bapx1-/-
mice. The expression domains of both genes correlate well with the phenotypes
observed in the knockout mice. Therefore, the cells destined to contribute to
the tympanic ring appear to be identified early in embryogenesis by the
expression domains of these two genes. Bapx1 and Gsc act in
concert to specify the thickened anterior end and thus are required for full
structural composition of the tympanic ring. Our observations show clearly
that in the branchial arch, Bapx1 and Gsc are regulated
independently in the region of overlap as deficiencies of either gene do not
affect the expression of the other. In addition, Bapx1 does not
regulate the expression of several other genes known to be involved in related
aspects of middle ear development, including Prx1, Msx1 and
Hand2.
Gsc has both an inductive and a cell autonomous role in the formation of the tympanic ring. Expression in the EAM, the epithelium adjacent to the tympanic mesenchyme, is required for condensation; whereas chondrogenesis is cell autonomous (Rivera-Pérez et al., 1999). Bapx1 in the gonium and tympanic ring, however, appears to be required from an early stage of specification, as no signs of condensation or chondrogenesis are apparent.
Evolution of the mammalian middle ear
The evolution of the middle ear in tetrapods is well documented in the
fossil record. The role of Bapx1 in the fish jaw suggested a
corresponding role in the middle ear of mammals. As anticipated,
Bapx1 does have a role in the skeletal pattern of the middle ear;
however, it does not directly correspond to that predicted by the zebrafish
data. To attempt to understand the molecular differences that may have
occurred in the transition to the middle ear, Bapx1 expression was
analysed in craniofacial morphogenesis in the developing chick. The facial
skeletal pattern in chick is an amenable experimental system related to the
transitional reptilian species important in the jaw/middle ear transformation
(Allin, 1975). Such analysis
enabled the comparison of expression patterns in relation to gross
evolutionary changes in morphology. Bapx1 expression in the first
branchial arch of vertebrates from fish to mammals reflects a continual role
for the gene in craniofacial evolution. Theory holds that evolution of
structure is founded on changes in gene regulatory systems. Hence,
Bapx1 is likely to hold a strategic position in a regulatory network
important in evolutionary change. Information from three vertebrate classes
enabled an examination of the hypothesis that changes in regulatory networks
are important in the morphological transformations.
Comparing the expression patterns of Bapx1 and Gsc, the spatial organisation in the early branchial arch was indistinguishable in chick and mouse. At the initial stage of expression, Gsc and Bapx1 overlap in the distal region of the mandibular arch in both mouse and chick. By E15.5, Bapx1 is expression is associated with the malleus and incus, the exact pattern depending on the part of the ossicle observed. Strong expression is detected between the malleus and incus at the site of articulation. At a similar developmental stage in chick (E7.5), Bapx1 is expressed in a comparable pattern, in between the quadrate and articular. Thus, even though the skeletal elements are distinct between species, similar characteristic Bapx1 and Gsc spatial patterns are established in chick and mouse. In the midst of gross morphogenetic and functional changes, there is a continuity of spatial expression. Hence, the basis for patterning of the mandibular arch is unmodified in the transition to the middle ear.
Gdf5 and Gdf6 are also expressed in the region surrounding the incus and malleus in mouse, and articular and quadrate in chick. This pattern overlaps with the expression of Bapx1 in both species. In fish, Gdf5 expression is dependent on Bapx1, whereas in mouse, the Gdf expression is unaffected in the Bapx1-/- mutant. Thus, several changes of the Gdf family members have occurred. First, a second GDF family member, Gdf6, is expressed in the mouse in a comparable pattern of Gdf5; and second, the expression of neither Gdf gene is controlled by Bapx1. Although Bapx1 has an overall role in the size of the malleus, the formation of the joint with the incus is independent of Bapx1 expression.
We suggest that Bapx1 spatial expression in branchial arch derivatives is continuous in vertebrate evolution; however, pertinent to the morphological transition are developmental processes downstream of the initial patterning events. Regulatory changes have occurred but these are to target genes of the regulatory network. Lack of control of Gdf expression by Bapx1 in mouse may reflect a number of downstream alterations that have occurred in the transition to middle ear ossicles and is a likely explanation for the differences in joint formation between mouse and fish. We suggest that the regulatory network involving the spatial expression of Bapx1 in vertebrate species is not detectably altered, but that the crucial evolutionary changes are downstream of the transcription factor and these are central to realisation of the species specific phenotypes.
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ACKNOWLEDGMENTS |
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Footnotes |
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* These authors have contributed equally to the work
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REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Akazawa, H., Komuro, I., Sugitani, Y., Yazaki, Y., Nagai, R. and
Nodal, T. (2000). Targeted disruption of the homeobox
transcription factor Bapx1 results in lethal skeletal dysplasia with
asplenia and gastroduodenal malformation. Genes Cells
5, 499-513.
Allin, E. F. (1975). Evolution of the mammalian middle ear. J. Morphol. 147,403 -437.[Medline]
Allin, E. F. and Hopson, J. A. (1992). Evolution of the auditory system in Synapsida ('mammal-like reptiles' and primitive mammals) as seen in the fossil record. In The Evolutionary Biology of Hearing (ed. D. B. Webster, R. R. Fay, A. N. Popper), pp. 587-614. New York: Springer-Verlag.
De Beer, G. (1937). The Development of the Vertebrate Skull. Chicago. IL: University of Chicago Press.
Depew, M. J., Tucker, A. S. and Sharpe, P. T. (2002). Mouse development-Patterning, Morphogenesis and Organogenesis. In Craniofacial Development (ed. J. Rossant and P. Tam), Chapter 19. London, Academic Press.
Ducy, P., Zhang, R., Geoffroy, V., Ridall, A. L. and Karsenty, G. (1997). Osf2/Cbfa1: a transcriptional activator of osteoblast differentiation. Cell 89,747 -754.[Medline]
Fleischer, G. (1978). Evolutionary principles of the mammalian middle ear. Adv. Anat. Embry. Cell Biol. 55,1 -70.
Gaunt, S. J., Blum, M., de Robertis, E. M.
(1993). Expression of the mouse goosecoid gene during
mid-embryogenesis may mark mesenchymal cell lineages in the developing head,
limbs and body wall. Development
117,769
-778
Gaupp, E. (1911). Beiträge zur Kenntnis des Unterkiefers der Wirbeltiere I; der Proc. Anterior (Folii) des Hammers der Säuger und das Gonial der Nichtsäuger. Anat. Anz. 39,97 -135.
Goodrich, E. S. (1986). Studies on the Structure and Development of Vertebrates. Chicago, IL. University of Chicago Press.
Grandien, K. and Sommer, R. J. (2001).
Functional comparison of the nematode Hox gene lin-39 in C. elegans and P.
pacificus reveals evolutionary conservation of protein function despite
divergence of primary sequences. Genes Dev.
15,2161
-2172.
Grenier, J. K. and Carroll, S. B. (2000).
Functional evolution of the Ultrabithorax protein. Proc. Natl.
Acad. Sci. USA 97,704
-709.
Hamburger, V. and Hamilton, H. L. (1951). A series of normal stages in the development of the chick embryo. J. Morphol. 88,49 -92.
Henson, O. W. (1974). Comparative anatomy of the middle ear. In Handbook of Sensory Physiology, Vol. V/1 (ed. W. D. Keidel and W. D. Neff), pp.81 -85. Berlin, Heidelberg, New York: Springer-Verlag.
Herbrand, H., Pabst, O., Hill, R. and Arnold, H. H. (2002). Transcription factors Nkx3.1 and Nkx3.2 (Bapx1) play an overlapping role in sclerotomal development of the mouse. Mech. Dev. 117,217 -224.[CrossRef][Medline]
Huangff, M. and Saunders, J. C. (1983). Auditory development in the mouse: structural maturation of the middle ear. J. Morphol. 176,249 -259.[Medline]
Kalatzis, V., Sahly, I., El-Amraoui, A. and Petit, C. (1998). Eya1 expression in the developing ear and kidney: Towards the understanding of the pathogenesis of Branchio-Oto-Renal syndrome. Dev. Dyn. 213,486 -499.[CrossRef][Medline]
Kessel, M. and Gruss, P. (1991). Homeotic transformations of murine vertebrae and concomitant alteration of Hox codes induced by retinoic acid. Cell 67, 89-104.[Medline]
Kim, Y. and Nirenberg, M. (1989). Drosophila NK-homeobox genes. Proc. Natl. Acad. Sci. USA 86,7716 -7720.[Abstract]
Komori, T., Yagi, H., Nomura, S., Yamaguchi, A., Sasaki, K., Deguchi, K., Shimizu, Y., Bronson, R. T., Gao, Y. H., Inada, M. et al. (1997). Targeted disruption of Cbfa1 results in a complete lack of bone formation owing to maturational arrest of osteoblasts. Cell 89,755 -764.[Medline]
Kuratani, S., Satakata, I., Blum, M., Komatsu, Y., Haraguchi, R., Nakamura, S., Suzuki, K., Kosai, K., Maas, R. and Yamada, G. (1999). Middle ear defects associated with the double knockout mutation of murine goosecoid and Msx1 genes. Cell. Mol. Biol. 45,589 -599.
Lettice, L. A., Purdie, L. A., Carlson, G. J., Kilanowski, F.,
Dorin, J. and Hill, R. E. (1999). The mouse bagpipe gene
controls development of axial skeleton, skull, and spleen. Proc.
Natl. Acad. Sci. USA 96,9695
-9700.
Mallo, M. (1997). Retinoic acid disturbs mouse middle ear development in a stage-dependent fashion. Dev. Biol. 184,175 -186.[CrossRef][Medline]
Mallo, M. (2001). Formation of the middle ear: recent progress on the developmental and molecular mechanisms. Dev. Biol. 231,410 -419.[CrossRef][Medline]
Mallo, M. and Gridley, T. (1996). Development
of the mammalian ear: coordinate regulation of formation of the tympanic ring
and the external acoustic meatus. Development
122,173
-179.
Martin, J. F., Bradley, A. and Olson, E. N. (1995). The paired-like homeo box gene Mhox is required for early events of skeletogenesis in multiple lineages. Genes Dev. 9,1237 -1249.[Abstract]
Miller, C. T., Yelon, D., Stainier, D. Y. R. and Kimmel, C.
B. (2003). Two endothelin 1 effectors,
hand2 and bapx1, pattern ventral pharyngeal cartilage and
the jaw joint. Development
130,1353
-1365
Newman, C. S., Grow, M. W., Cleaver, O., Chia, F. and Krieg, P. (1997). Xbap, a vertebrate gene related to bagpipe, is expressed in developing craniofacial structures and in anterior gut muscle. Dev. Biol. 181,223 -233.[CrossRef][Medline]
Reichert, C. (1837). Über die Visceralbogen der Wirbeltiere im allgemeinen und deren Metamophose bei den Vögelm und Säugetieren. Arch. Anat. Physiol.120 -222.
Rivera-Pérez, J. A., Mallo, M., Gendron-Maguire, M.,
Gridley, T. and Behringer, R. R. (1995). goosecoid is not an
essential component of the mouse gastrula organizer but is required for
craniofacial and rib development. Development
121,3005
-3012
Romanoff, A. L. (1960). The Avian Embryo: Structural and Functional Development. New York: Macmillan.
Rosowski, J. J. (1992). Hearing in transitional mammals. In The Evolutionary Biology of Hearing (ed. D. B. Webster, R. R. Fay and A. N. Popper), pp.615 -631. New York: Springer-Verlag.
Satokata, I. and Maas, R. (1994). Msx1 deficient mice exhibit cleft palate and abnormalities of craniofacial and tooth development. Nat. Genet. 6, 348-356.[Medline]
Schilling, T. and Kimmel, C. (1997). Musculoskeletal patterning in the pharyngeal segments of the zebrafish embryo. Development 120,483 -494.
Settle, S. H., Jr, Rountree, R. B., Sinha, A., Thacker, A., Higgins, K. and David, M. and Kingsley, D. M. (2003). Multiple joint and skeletal patterning defects caused by single and double mutations in the mouse Gdf6 and Gdf5 genes. Dev. Biol. 254,116 -130.[CrossRef][Medline]
Sharpe, J., Ahlgren, U., Perry, P., Hill, B., Ross, A.,
Hecksher-Sorensen, J., Baldock, R. and Davidson, D. (2002).
Optical projection tomography as a tool for 3D microscopy and gene expression
studies. Science 296,541
-545.
Storm, E. E., Huynh, T. V., Copeland, N. G., Jenkins, N. A., Kingsley, D. M. and Lee, S.-J. (1994). Limb alterations in brachypodism mice due to mutations in a new member of the Tgf-beta superfamily. Nature 368,639 -643.[CrossRef][Medline]
Storm, E. E. and Kingsley, D. M. (1999). Gdf5 coordinates bone and joint formation during digit development. Dev. Biol. 209,11 -27.[CrossRef][Medline]
Tanaka, M., Lyons, G. E. and Izumo, S. (1999). Expression of the Nkx3.1 homeobox gene during pre and postnatal development. Mech. Dev. 85,179 -182.[CrossRef][Medline]
Tribioli, C. and Lufkin, T. (1997). Molecular cloning, chromosomal mapping and developmental expression of Bapx1, a novel human homeobox-containing gene homologous to Drosophila bagpipe. Gene 203,225 -233.[CrossRef][Medline]
Tribioli, C. and Lufkin, T. (1999). The murine
Bapx1 homeobox gene plays a critical role in embryonic development of
the axial skeleton and spleen. Development
126,5699
-5711.
Tribioli, C., Frasch, M. and Lufkin, T. (1997). Bapx1: an evolutionary conserved homologue of the Drosophila bagpipe homeobox gene is expressed in splanchnic mesoderm and the embryonic skeleton. Mech. Dev. 65,145 -162.[CrossRef][Medline]
Trumpp, A., Depew, M. J., Rubenstein, J. L. R., Bishop, M. J.
and Martin, G. R. (1999). Cre-mediated gene inactivation
demonstrates that FGF8 is required for cell survival and patterning of the
first branchial arch. Genes Dev.
13,3136
-3148.
Tucker, A. S., Al Khamis, A., Ferguson, C. A., Bach, I.,
Rosenfeld, M. G. and Sharpe, P. T. (1999). Conserved
regulation of mesenchymal gene expression by Fgf-8 in face and limb
development. Development
126,221
-228.
Voit, M. (1924). Uber das Goniale am Unterkiefer der Vogel. Z. Morphol. Anthrop. 24, 75-82.
Wilkinson, D. (1992). Whole mount in situ hybridization of vertebrate embryos. In In Situ Hybridization: A Practical Approach (ed. D. Wilkinson), pp.75 -83. Oxford: IRL Press at Oxford University Press.
Wilson, J. and Tucker, A. S. (2004). Fgf and Bmp signals repress the expression of Bapx1 in the mandibular mesenchyme and control the position of the developing jaw joint. Dev. Biol. (in press).
Xu, P-X., Adams, J., Peters, H., Brown, M. C., Heaney, S. and Maas, R. (1999). Eya-1 deficient mice lack ears and kidneys and show abnormal apoptosis of organ primordia. Nature 23,113 -117.[CrossRef]
Yamada, G., Mansouri1, A., Torres, M., Stuart, E. T., Blum, M.,
Schultz, M., de Robertis, E. M. and Gruss, P. (1995).
Targeted mutation of the murine goosecoid gene results in craniofacial defects
and neonatal death. Development
121,2917
-2922.