Department of Biological Sciences, University of Pittsburgh, Pittsburgh,
PA 15260, USA
* Present address: UC Davis School of Veterinary Medicine, Davis, CA 95616,
USA
Author for correspondence (e-mail:
dlc7{at}pitt.edu)
Accepted 20 December 2002
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SUMMARY |
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Key words: Mouse, Somitogenesis, Segmentation, T-box, Tbx6, Dll1, rib-vertebrae
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INTRODUCTION |
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Notch signaling is essential for establishing and maintaining somite
boundaries by setting up differences between R-C halves of somites; this
occurs within the anterior presomitic mesoderm (PSM), prior to somite
formation. Mutations in various components of the Notch signaling pathway
disrupt the process of somite segmentation and the latter process of
resegmentation (reviewed by Pourquie and
Kusumi, 2001). For example, mutations in the human DLL3,
a Notch ligand, results in spondylocostal dysostosis, which is characterized
by hemivertebrae, rib fusions and deletions that together lead to short
stature in the affected individuals (Bulman
et al., 2000
). Improper subdivision of the somites into R-C halves
results most noticeably in the formation of fused ribs and vertebrae. It may
also result in defective patterning of other tissues, in particular the
peripheral nervous system (PNS). The neural crest-derived spinal ganglia
migrate through the rostral halves of the somites, thus giving the PNS a
segmental arrangement, so that disruption of R-C somite patterning disrupts
the segmental arrangement of the PNS (reviewed by
Bronner-Fraser, 2000
).
The mouse T-box transcription factor Tbx6 is expressed in the
primitive streak and PSM, and is downregulated as the paraxial mesoderm
segments to form the somite (Chapman et
al., 1996). Gene targeting studies in mice revealed that Tbx6 is
necessary for the formation of paraxial mesoderm posterior to the forelimb
bud, which is replaced by ectopic neural tubes in the Tbx6-null
mutant embryos (Chapman and Papaioannou,
1998
). Primitive streak markers T, Wnt3a and
Fgf8 are all expressed in the Tbx6-null mutant, while PSM
markers such as Notch1 and paraxis are not, suggesting that
differentiation of primitive streak into PSM is blocked
(Chapman and Papaioannou, 1998
)
(D.L.C., unpublished).
The Notch ligand, delta-like 1 (Dll1) is expressed in the
primitive streak and PSM, in a domain that overlaps with Tbx6
(Bettenhausen et al., 1995;
Chapman et al., 1996
). Like
other genes that are normally expressed in the PSM, Dll1 is not
expressed in the Tbx6-null mutant embryos
(Chapman and Papaioannou,
1998
), suggesting that it might be a target of Tbx6.
Alternatively, its expression may be lost simply because the PSM is not
present in the mutant. Herein, however, we present data that supports Tbx6
functioning through Dll1 in patterning the somites. First, we show that
reduction of Tbx6 expression in embryos below heterozygous levels
leads to fusions of the ribs and vertebrae, a characteristic shared by Notch
signaling mutants. We further show that these malformed vertebrae are due to
improper patterning of the somites along their R-C axis. Second, we
demonstrate, not only that Tbx6 genetically interacts with the
classic mouse mutant rib-vertebrae (rv), but also that
rv is a hypomorphic mutation in Tbx6. Finally, we show that
Tbx6 genetically interacts with Dll1, and that this,
together with the absence of Dll1 expression in Tbx6-null
mutant embryos, suggest that Tbx6 is upstream of Dll1 in the pathway
leading to somite formation and patterning.
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MATERIALS AND METHODS |
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Mice
The Tbx6tm1Pa mice have previously been described and
are maintained on a mixed C57Bl6/J 129Sv/Ev genetic background
(Chapman and Papaioannou,
1998). Dll1tm1Gos and rib-vertebrae
(rv) mutant mice were obtained from Jackson Laboratories and are
maintained on a C57Bl6/J background
(Beckers et al., 2000b
;
Hrabe de Angelis et al., 1997
;
Nacke et al., 2000
). The two
Tbx6 transgenic lines (Tg46 and Tg130) are
maintained on both FVB/N and a mixed genetic background. Genotyping of the
Tbx6 and Dll1 mutant mice was performed as previously
described (Chapman and Papaioannou,
1998
; Hrabe de Angelis et al.,
1997
). The rv allele was genotyped by Southern blot
analysis using a Tbx6-specific genomic probe that detects a
polymorphism between the Tbx6 wild-type and rv mutant
alleles. Similarly the Tbx6 transgene was detected using a
Tbx6-specific probe by Southern blot analysis.
PCR and cloning of rv lesion
Genomic DNA from rv/rv and wild-type C57Bl6/J mice was used for
PCR using the following primers designed from sequences obtained at Ensembl
Mouse Genome Server: (forward) 5'-CATTCCCAAAACCCCATTGC -3',
(reverse) 5'-TGCCCCTTCACTCTCTCCATC -3'; and (forward)
5'-GTGTAGTTGAAATGTTCTCGGCG -3', (reverse)
5'-CACAGTTCCTGGTTCTCCAAGC -3'. The PCR products were cloned and
sequenced as described above.
Whole-mount in situ hybridization
Whole-mount in situ hybridization was performed as previously described by
Wilkinson (Wilkinson, 1992)
using antisense riboprobes for Tbx6, Tbx18, Dll1, Dll3. Mesp2, Uncx4.1,
Cer1, neurofilament L and myogenin. Hybridization and washes were
performed at 63°C.
X-gal staining
Embryos were dissected at the indicated times and stained for
ß-galactosidase activity with X-gal as previously described
(Ciruna et al., 1997).
Skeletal preparations
Skeletons from embryos dissected at E14.5 and E15.5 were stained with
Alcian Blue (staining the cartilage) and Alizarin Red (staining the bone) as
described by Hogan et al. (Hogan et al.,
1994), except that staining was performed simultaneously at
37°C. After staining, embryos were cleared in 1% KOH and stored in 1%
KOH:glycerol (90:10).
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RESULTS |
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Rostrocaudal patterning of the somites is disrupted in Tbx6
transgenic embryos
Through our attempts to rescue the Tbx6 mutant using a transgene
that expresses Tbx6 at low levels, we have established a series of
Tbx6 phenotypes characterized by rib and vertebral fusions, which
suggests improper patterning of the somites. To explore this possibility more
carefully, we performed whole-mount in situ hybridization using markers of
somite R-C patterning on Tg46 rescued embryos and their littermates
derived from crosses of Tbx6tm1Pa/+ Tg46/+ x
Tbx6tm1Pa/+ mice.
The paired homeobox gene Uncx4.1 is normally expressed in the
caudal halves of somites (Mansouri et al.,
1997; Neidhardt et al.,
1997
) (Fig. 3A). In
the Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ embryo,
Uncx4.1 transcripts are detected throughout the somitic region with
no apparent stripes (Fig.
3B,C). In some of these embryos, Uncx4.1 appears as
darker stripes in the most recently formed somites; however, these stripes are
found amidst uniform, low-level Uncx4.1 expression
(Fig. 3D). Thus, somites in
these partially rescued Tbx6 mutants appear to be caudalized.
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Mouse cerberus-related gene 1, Cer1, is expressed as two stripes
in the anterior PSM and in the rostral region of the most recently formed
somite (Biben et al., 1998;
Shawlot et al., 2000
)
(Fig. 3F). As expected,
Cer1 is not expressed in the Tbx6-null mutant (data not
shown). Cer1 is expressed in the PSM of Tg46 rescued
embryos, but levels are reduced compared with control embryos, and no
expression is detected in the newly formed somite
(Fig. 3F). This result also
suggests that rostral somite compartment identity of the PSM is initially
observed; however, it is not maintained in these embryos.
Neural crest-derived spinal ganglia migrate through the rostral half of the
sclerotome, therefore loss or improper formation of this somite compartment
will lead to alterations in the appearance of the spinal ganglia. To
investigate this, we examined the expression of neurofilament L, which is
segmentally expressed in the spinal ganglia along the anteroposterior (AP)
axis of normal embryos (Lewis and Cowan,
1985) (Fig. 3G).
The rostral compartment of the somites is clearly disrupted in the
Tg46 rescued embryos as shown by the expression pattern of
neurofilament L. In contrast to the segmented appearance of the neurofilament
L-expressing spinal ganglia in the normal embryo, the segments are
disorganized and often joined in rescued embryos
(Fig. 3H).
The T-box gene Tbx18 is expressed in the rostral region of somites
along the AP axis of the embryo, in addition to the heart and limb buds
(Kraus et al., 2001)
(Fig. 3I,K). A range of
expression patterns was observed for this gene in the Tg46 rescued
embryos. Some rescued embryos failed to express Tbx18 in the somites
(Fig. 3J), while other embryos
expressed reduced levels of Tbx18 transcripts in the rostral region
of the newly formed somites, but this expression was not maintained in more
anterior somites (Fig. 3L). In
between these two extremes were embryos that expressed Tbx18 at
reduced levels and in smaller domains in the most newly formed somites.
However, it was no longer expressed in more anterior regions of the embryo
(data not shown). Altogether these marker gene studies show that R-C somite
patterning is clearly disrupted in the Tg46 rescued mutant embryos,
with somites losing their rostral compartment identity.
Expression of Dll1, but not Dll3, is reduced in
Tbx6 transgenic embryos
The defect in R-C somite patterning in the
Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ embryos suggests
that expression of one or more components of the Notch signaling pathway is
disrupted in these embryos. Two Notch ligands, Dll1 and Dll3, are known to be
involved in R-C somite patterning
(Barrantes et al., 1999;
Dunwoodie et al., 2002
).
Dll3 is normally expressed in the primitive streak, PSM and later in
the rostral somite compartment of the two most recently formed somites
(Dunwoodie et al., 1997
). In
contrast to most PSM expressed genes, including Dll1, Dll3 is
expressed in the Tbx6-null mutant and its expression in the tailbud
of the Tg46 rescued embryos is comparable with controls
(Fig. 3M). Dll3
expression in the rostral regions of the newly formed somites of the
Tg46 rescued embryos is not apparent; however, expression in this
domain was not seen in control embryos either
(Fig. 3M).
Dll1 is normally expressed in the primitive streak and PSM, then
localizing to the caudal halves of somites
(Bettenhausen et al., 1995)
(Fig. 3N,Q). Although
Dll1 is not expressed in the tail mesoderm of the Tbx6-null
embryo (Chapman and Papaioannou,
1998
), Dll1 expression is restored in the
Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ embryos, with
transcripts present in the PSM with a slight upregulation in the somite that
is next to form (Fig. 3O,P).
Dll1 expression levels, however, were reduced in the Tg46
rescued embryos compared with Tbx6 heterozygotes (compare
Fig. 3N,Q with 3O,P,R,S).
Similar to ß-galactosidase staining, Dll1 transcripts localize
to the dorsal region of the expanded tail region and are absent from more
ventral areas (Fig. 3R,S). Low
levels of Dll1 expression in the caudal halves of somites can be seen
in both the Tbx6 heterozygotes and in the partially rescued mutants
(Fig. 3N-S). In addition to
this mesoderm expression, Dll1 is also expressed in neural tissue
(Bettenhausen et al., 1995
).
Dll1 expression is detected in the flanks of some Tg46
rescued embryos, marking the ectopic neural tissue, which still forms in the
paraxial region of some of these mutants despite the presence of paraxial
somites (Fig. 3P).
We have also examined expression of other Notch signaling pathway genes in the Tbx6 nulls and Tg46 rescued embryos, specifically Notch1 and lunatic fringe (Lfng). Although neither gene is expressed in the Tbx6-null embryos, the expression of both is restored in the PSM of Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ embryos; however, this expression is reduced compared with normal embryos (data not shown). The dynamic expression of Lfng in the PSM was observed in the Tg46 rescued embryos, but transcripts were present at reduced levels. The reductions in expression of the Notch signaling pathway genes Dll1, Notch1 and Lfng in the Tg46 rescued embryos compared with normal littermates could individually or together account for the R-C somitic defects observed.
Tbx6 genetically interacts with rib-vertebrae
The phenotype of the partially rescued Tbx6 mutant embryos was
reminiscent of the of the rib-vertebrae (rv) mutant
phenotype. rv is a spontaneous recessive mutation characterized by
fusions of ribs and vertebrae, shortened trunk, kinked tail, formation of a
single kidney, and reduced fertility
(Beckers et al., 2000b;
Nacke et al., 2000
;
Theiler and Varnum, 1985
).
rv homozygous embryos are recognized by an enlarged tailbud, which
later in development has multiple outgrowths [which are characteristics shared
with the Tg46 rescued embryos
(Fig. 2D)], and by caudal
duplications of the neural tube [which is a characteristic shared with the
Tbx6-null mouse (Chapman and
Papaioannou, 1998
; Theiler and
Varnum, 1985
)]. Interestingly, rv had been mapped to
position 62 cM on mouse chromosome 7
(Beckers et al., 2000b
), which
was close to the mapped location of Tbx6 at position 61 cM
(Chapman et al., 1996
).
Altogether, these data suggest that Tbx6 is the gene affected in the
rv mutation. To determine whether Tbx6 interacts genetically
with rv, embryos were dissected from Tbx6tm1Pa/+
x rv/rv crosses at E15.5 and their skeletons were stained with
Alcian Blue and Alizarin Red (Fig.
4A-E). At the level of gross morphology, compound heterozygous
embryos can be distinguished by their short stature and short tails
(Fig. 4B), similar to the
Tg46 rescued embryos. The genetic interaction between these two
mutations is further illustrated by skeletal preparations of these embryos,
which clearly show fusions of ribs and vertebrae along the entire AP axis
(Fig. 4D,E).
Tbx6tm1Pa/+ rv/+ pups are either resorbed during
development or are found dead at birth. Examination of Tbx6
expression in rv/rv and rv/+ embryos revealed that although
Tbx6 was expressed in the correct spatial pattern, transcripts were
present at lower levels in the tailbuds of rv/rv compared with
rv/+ (Fig. 4F). In
addition, Tbx6 expression is reduced in the compound mutant embryos
compared with Tbx6tm1Pa/+ (data not shown). These results
indicate that Tbx6 expression is directly affected in the rv
mutant, and that the genetic interaction observed between the two mutants is
probably due to reduced levels of Tbx6 expression.
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The defect in somite formation in rv/rv mice is due to disruption
of R-C patterning of the somites and this ultimately leads to the fusions of
ribs and vertebrae (Beckers et al.,
2000b; Nacke et al.,
2000
). rv genetically interacts with Dll1
(Beckers et al., 2000b
;
Nacke et al., 2000
). As we
have identified rv as an allele of Tbx6, the targeted mutant
allele of Tbx6 (Tbx6tm1Pa) should also interact
genetically with Dll1. We confirmed this by generating
Tbx6tm1Pa/+ Dll1tm1Gos/+ embryos and mice.
Examination of the Tbx6tm1Pa/+ Dll1tm1Gos/+
embryonic skeletons revealed a variety of vertebral defects, including fusions
of the neural arches in cervical vertebrae, missing regions of ribs and
abnormally formed vertebrae along the AP axis
(Fig. 4I-K).
Tbx6tm1Pa/+ Dll1tm1Gos/+ mice are born and can
be distinguished by their kinked tails (not shown). The phenotypes of these
compound heterozygotes were much milder than those seen for either the
Tg46 rescued embryos or the Tbx6tm1Pa/rv
mutant embryos described above.
Defects in myotome in Tbx6 hypomorphic embryos
Whether reduced Tbx6 expression in the embryo is achieved by
transgene rescue of the Tbx6-null or by a bona fide Tbx6
hypomorphic mutation, patterning of the myotome compartment of the somite is
also clearly disrupted. Marker gene analysis of the
Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ and
Tbx6tm1Pa/rv embryos revealed irregular patterning of the
myotome in these mutant embryos (Fig.
5). Myogenin is normally expressed in distinct stripes along the
AP axis, marking the myotome (Cheng et
al., 1992; Sassoon et al.,
1989
; Wright et al.,
1989
) (Fig. 5A,C).
In both the Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ and the
Tbx6tm1Pa/rv embryos, myogenin expression no longer has an
evenly spaced striped pattern, but instead has areas of fused adjacent
myotomes, as well as isolated pools of myogenin-expressing cells along the AP
axis (Fig. 5B,D). Defects in
these Tbx6 hypomorphs therefore affect multiple compartments of the
somite.
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DISCUSSION |
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In Tbx6-null mutant embryos, somites posterior to the forelimb bud fail to form, and mesodermal tissue in the tails of these mutants appears to be arrested at the primitive streak stage of differentiation. It was not clear from these studies, however, if Tbx6 played any role in patterning of the somites or if it was simply required for establishing the paraxial mesoderm fate. We show that embryos with Tbx6 expression levels intermediate between null homozygotes and heterozygotes develop somites but that these are incorrectly patterned along their R-C axis, which leads to abnormalities in somite-derived tissue, most notably the axial skeleton. These embryos were generated in two ways: first, by using a rescuing transgene containing the Tbx6-coding region and cis-regulatory elements in a Tbx6 null mutant; and, second, by identifying a previously known mutation, rv, as a hypomorphic mutation in Tbx6.
Disruption of R-C somite patterning in embryos with reduced
Tbx6 expression levels
Expression studies in embryos with reduced Tbx6 expression reveal
that the R-C division is clearly disrupted. In both the rv/rv and
Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ embryos, the caudal
somite compartment marker Uncx4.1 is expressed in a wider somite
domain compared with normal embryos, suggesting that somites in these embryos
have adopted a caudal phenotype (Fig.
3A-D) (Beckers et al.,
2000b). The degree of caudalization is correlated with the
severity of the phenotypes and hence with the level of Tbx6
expression. The low level of Tbx6 expression observed in the
Tg46 rescued embryos results in almost complete caudalization of the
somites (as assessed by Uncx4.1 expression) and to severe fusions of
the ribs and vertebrae along the entire AP axis of the embryo. In
rv/rv embryos, intermediate levels of Tbx6 (between the
Tg46 rescued embryos and Tbx6 heterozygotes) result in
somites with wider than normal domains of Uncx4.1 expression;
however, unstained regions in these somites were also observed
(Beckers et al., 2000b
).
Consequently, the rib and vertebral fusions were not as severe in the
rv/rv embryos compared with Tg46 rescued embryos
(Fig. 2I)
(Beckers et al., 2000b
).
Although somites in the Tg46 rescued embryos appear caudalized,
marker genes specific for the rostral compartment of the forming somite, such
as Mesp2, Cer1 and Tbx18, are still present. Mesp2
marks the rostral half of the forming somite and plays a role in specifying
rostral somite compartment identity, as paraxial mesoderm in the
Mesp2 homozygous mutant embryos becomes caudalized
(Saga et al., 1997
).
Mesp2 is still expressed in Tg46 rescued embryos and
suggests that rostral somite identity is initially established in these
embryos; however, it is eventually lost, based on Uncx4.1 expression.
This result is supported by the expression of another rostral somite marker,
Cer1. Cer1 is expressed in the anterior PSM of the Tg46
rescued embryos, but is expressed at low levels and is not expressed in a
third stripe in the most recently formed somite, again supporting an eventual
loss of rostral compartment identity. In this regard, our data for
Cer1 expression in the Tbx6tm1Pa/Tbx6tm1Pa
Tg46/+ embryos differs from that previously reported for the
rv/rv mutant embryos, where Cer1 expression was shown to be
absent (Beckers et al., 2000b
).
It is unclear why this difference was observed between the rv/rv and
Tbx6tm1Pa/Tbx6tm1Pa Tg46/+ embryos, especially
as the rostral marker Mesp2 was expressed in the rv/rv
embryos, indicating that the rostral compartment is formed. Further analysis
of rostral somite compartment identity revealed that Tbx18, which is
normally expressed in the rostral halves of somites along the AP axis of the
embryo, is either not expressed or is expressed at lower levels in the most
recently formed somites, and is lost in more anterior somites of the
Tg46 rescued embryos. The irregular R-C somite patterning is further
exemplified by neurofilament L expression in the neural crest-derived spinal
ganglia, which normally appear as discrete segments along the AP axis because
of their migration through the rostral halves of somites. In our Tg46
rescued embryos, neurofilament L expression appears disorganized with segments
fusing.
Segmentation of the PSM is believed to be dependent on these R-C
differences (Stern and Keynes,
1987). Clear somite boundaries appear to be established in embryos
with reduced levels of Tbx6, suggesting that some differences in R-C
compartments are initially established in these embryos. These R-C
differences, however, are not sufficiently maintained, as demonstrated by
Tbx18 and neurofilament L expression patterns. The failure to
establish firmly and/or maintain R-C differences lead to later defects in
resegmentation, which can account for the vertebral defects observed. Although
the myotome is not directly dependent on R-C patterning of the somite, the
myotome is affected in the Tbx6 hypomorphs. Our results suggest
therefore that the failure to maintain these R-C differences will indeed
affect other compartments of the somite, perhaps because of a failure to
maintain somite boundaries.
Dll1, a potential target of Tbx6
The disruption in somite patterning observed in embryos with reduced levels
of Tbx6 is similar to that of Notch signaling component mutants,
suggesting that the expression or activity of one or more genes involved in
this pathway is disrupted in the presomitic mesoderm of these embryos. The
Notch ligand Dll1 is an obvious candidate, because expression is
completely lost in the Tbx6-null embryo
(Chapman and Papaioannou,
1998). Dll1 expression is restored in Tg46
rescued embryos, but the level of Dll1 transcripts is clearly reduced
compared with that in Tbx6 heterozygotes. Furthermore, previous
studies with rv and our studies here with the Tbx6-null
allele have revealed a strong genetic interaction between Dll1 and
Tbx6: Tbx6tm1Pa/+ Dll1tm1Gos/+ embryos
have defects in rib and vertebral patterning, although this phenotype is much
weaker than rv/rv or Tbx6tm1Pa/Tbx6tm1Pa
Tg46/+. Thus, one likely cause for the defective somite patterning in
embryos with reduced levels of Tbx6 is a reduction in Dll1
expression. As demonstrated by gene targeting experiments, Dll1 is
required for proper R-C patterning of the somites and for epithelialization of
the somites (Hrabe de Angelis et al.,
1997
).
Dll3 expression in embryos with reduced Tbx6 expression
is indistinguishable from wild type, indicating that there is not a simple
reduction in expression of all presomitic-specific genes in these embryos.
Consistent with this, no genetic interaction between Dll3 and
rv was detected (Beckers et al.,
2000b). Notch1 and Lfng, which are also required
for somite formation and patterning, are expressed at reduced levels in the
PSM of these embryos, but in the correct spatial patterns. The reduced
expression levels of these genes might contribute to the observed phenotypes.
However, no genetic interaction between Notch1 and rv was
detected (Beckers et al.,
2000b
), again implicating Dll1 as the primary factor disrupted in
these embryos.
One problem with directly linking Tbx6 and Dll1 is that
the mutants appear to have opposite phenotypes in terms of their effects on
compartmentalization of the somites. In Dll1-null mutants, somites
appear to be rostralized (Barrantes et al.,
1999), in contrast to their caudalization in embryos with reduced
levels of Tbx6. At present, the reason for this is not clear. It is
likely that Tbx6 is regulating the expression of other genes besides
Dll1, as would be expected based on the differences in the
Tbx6 and Dll1 null phenotypes. Other likely candidates are
Notch1 and Lfng, the expression of which was restored in
embryos expressing lowered levels of Tbx6 compared with their lack of
expression in the homozygous null. The combined effects of lowered Dll1,
Notch1 and Lfng may account for the differences seen in
compartment phenotypes. A complete explanation awaits the identification of
these additional Tbx6 targets. Another explanation is that reduction rather
than complete loss of Dll1 may result in caudalization rather than
rostralization: this can only be tested directly with Dll1
hypomorphs.
Tbx6 is a transcription factor and thus could be directly activating
Dll1. The regulatory elements necessary for Dll1 expression
have been identified through transgenic studies
(Beckers et al., 2000a). We
have examined these regulatory elements for T-box-binding sites using the
binding site identified for T (Kispert and
Herrmann, 1993
); however, no palindromic or half sites were
identified. As the binding site for Tbx6 has not yet been identified, further
analysis along these lines awaits the identification of a Tbx6-specific
binding site.
Human disorders caused by Tbx6 mutations?
Mouse embryos that bear homozygous Tbx6-null mutations result in a
lethal phenotype by embryonic day 12.5
(Chapman and Papaioannou,
1998). It is likely that similar mutations in humans would also
lead to early embryonic lethal phenotypes that would probably appear as
spontaneous abortions. Mice heterozygous for the Tbx6 mutation are
normal and fertile, with no signs of skeletal or muscle abnormalities,
suggesting that humans heterozygous for Tbx6-null mutations would
presumably be normal. This paper identifies phenotypes that lie between these
two extremes, with hypomorphic levels of Tbx6 expression resulting in
R-C patterning defects in the somites. Thus far, no human syndromes that would
result from improper R-C somite patterning map to human TBX6 on
chromosome 16. However, several syndromes like Klippel-Feil and spondylocostal
and spondylothoracic dysostosis, which display a number of clinical defects
including rib and vertebral fusions, may in the future be linked to
hypomorphic mutations in TBX6 or to genes functioning in this
pathway.
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ACKNOWLEDGMENTS |
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