Division of Developmental Biology, Children's Hospital Medical Center,
3333 Burnet Avenue, Cincinnati, OH 45224, USA
*
Author for correspondence (e-mail:
steve.potter{at}chmcc.org
)
Accepted 30 May 2001
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SUMMARY |
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Key words: Hox, Hoxa13, Hoxa11, Homeobox, Functional specificity, Abd-B, Downstream targets, Mouse
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INTRODUCTION |
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The evidence arguing for functional equivalence of paralogous mammalian Hox
genes is particularly strong. Paralogs reside at equivalent positions within
different Hox clusters and are derived by evolutionary gene duplication from a
single ancestral Hox gene. They generally encode very similar homeodomains,
and show overlapping gene expression patterns. Gene targeting experiments have
shown striking functional redundancy for paralogs. For example, mice mutant
for either Hoxa11 or Hoxd11 show no kidney defects and only
mild limb defects (Small and Potter,
1993; Davis and Capecchi,
1994
). But mice mutant for
both these paralogs show absent or rudimentary kidneys, and have the ulna and
radius of the forearm reduced to less than one tenth of normal size (Davis et
al., 1995
). In addition, a
Hoxd11-expressing transgene has been shown to be able to rescue
Hoxa11 loss of function (Zakany et al.,
1996
). Furthermore, complete
coding sequence exchanges between the Hoxa3 and Hoxd3
paralogs have indicated that their proteins carry out identical biological
functions (Greer et al.,
2000
). These results have been
interpreted to support the functional equivalence model (Duboule,
2000
).
Although it has been argued by some that the Drosophila paradigm
may not apply to mammalian Hox gene function, it is nevertheless useful to
consider what has been learned from that system. In Drosophila there
is one homeotic complex (HomC) of genes, which represents a single split
cluster. Ectopic expression experiments have shown that different HomC genes
can induce very distinct developmental destinies. For example, misexpression
of Antennapedia can cause imaginal disc cells that would normally
form antennae to instead give rise to legs, protruding from the head
(Schneuwly et al., 1987). And
misexpression of Ultrabithorax can result in the homeotic
transformation of wing into haltere (Lewis,
1982
). The combinatorial code
expression of HomC genes has been proposed to determine segment identity
(Lewis, 1978
). The apparently
distinct functional specificities of different HomC genes have several
sources. First, the encoded homeodomains are not functionally equivalent.
Homeobox swap experiments in Drosophila show that developmental
function often tracks with the homeobox, suggesting the presence of in vivo
target sequence binding specificity not detected by in vitro DNA-binding
assays (Kuziora and McGinnis,
1989
; Gibson et al.,
1990
; Mann and Hogness,
1990
). Second, target
specificity is influenced by co-factor interactions. Different Exd-HomC
protein heterodimers, for example, have distinct DNA-binding specificites,
even when measured in vitro (Chan et al.,
1994
; Mann and Chan,
1996
), and there is evidence
for similar gains in specificity in mammalian systems (Popperl et al.,
1995
; Chan et al.,
1997
). Third, different HomC
proteins may possess activation or repression domains, which can determine
developmental function by their distinct impacts on the same target gene
expression. Fourth, according to the activity regulation model, binding sites
for specific combinations of co-factors in a given promoter would confer
activation or repression effects (Li et al.,
1999
). This is consistent with
the observation that individual HomC proteins are often able to repress some
target genes and activate others, as measured by both genetic and transfection
assays (Vachon et al., 1992
;
Capovilla et al., 1994
; Saffman
and Krasnow, 1994
). In
summary, a number of mechanisms exist in Drosophila for providing
individual HomC proteins with functional specificity. It would seem reasonable
to suppose that similar molecular processes might be used in mammals.
Nevertheless, even in Drosophila there is considerable evidence
for functional overlap of HomC genes. For example, misexpression of Ubx,
Abd-A or Abd-B can cause cells that would normally form wing to
form haltere instead (Casares et al.,
1996). This is particularly
informative, as Abd-A and Abd-B are normally only involved
in development of the abdomen, which has no wings or halteres. It has also
been shown that a hybrid Ubx-VP16, with enhanced transcription
activation function, mimics Antp in developmental specificity,
presumably by regulating the same set of downstream targets (Li and McGinnis,
1999
). Furthermore, several
promoter analysis studies suggest that the HomC genes regulate overlapping
sets of downstream target genes (Manak et al.,
1994
; Mastick et al.,
1995
). These observations
suggest that the differences in function between the Drosophila HomC
and mammalian Hox genes are less pronounced than proposed by the mammalian Hox
functional equivalence model. Perhaps the Drosophila HomC genes are
not each functionally unique, and perhaps the mammalian Hox genes are not all
functionally the same.
To address multiple issues of homeodomain function, including the question of functional specificity, we performed a mammalian homeobox swap experiment. The homeobox of the Hoxa11 gene was precisely replaced with that of the Hoxa13 gene. The Hoxa11 and Hoxa13 genes are closely related Abd-B type genes. In a broad sense they can be considered paralogs, as they are derived from a common ancestral Abd-B Hox gene. The functional equivalence model proposes that these two genes have identical or functionally equivalent downstream targets, and therefore predicts that the swapped allele would have wild-type function. This was indeed observed in the developing kidneys, male reproductive tract and axial skeleton. Striking mutant phenotypes were seen, however, in the limbs and female reproductive tract. Of particular note, in mice with the swapped allele, the uterus underwent a homeotic transformation towards cervix/vagina, as determined by both histology and gene chip analysis of gene expression profiles. This homeotic transformation indicates a patterning function for Hox genes. The altered gene expression profiles identify candidate Hoxa13 downstream targets. The results indicate that the Hoxa11- and Hoxa13-encoded homeodomains are not functionally equivalent, and that in some developing tissues the Hoxa11 allele with a swapped Hoxa13 homeobox assumed Hoxa13 developmental function.
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MATERIALS AND METHODS |
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The released 9 kb NheI segment was subcloned into a modified pBS, with the multiple cloning site XhoI and BSTXI sites removed, giving construct II. This clone had unique XhoI and BSTXI sites flanking the A11 second exon with the homeobox. This XhoI-BSTXI segment was subcloned into pBS, giving the A11 (XhoI-BstXI) vector.
Two new restriction sites, HincII and PstI were
introduced at the junction regions of the A11 homeobox of the
XhoI-BSTXI segment by PCR mutagenesis (Bi and Stambrook,
1998). The primers used
were:
Lowercase letters represent restriction enzyme sites, underlines indicate silent mutation sites, bold letters represent homeobox region and parenthesized letters represent sequence not included in the primer sequence.
The A13 homeobox was PCR amplified from strain 129 DNA using the following primers.
The A13 homeobox PCR product was digested with HincII and PstI and subcloned into the A11 (XhoI-BSTXI) vector, also cut with HincII and PstI, replacing the A11 homeobox. This XhoI-BSTXI segment was then subcloned into XhoI, BSTXI cleaved construct II, and the 9 kb NheI segment of the modified construct II was then subcloned back into NheI cut construct I, making the final targeting construct. The construct was confirmed by DNA sequencing.
Gene targeting
The targeting constructs were introduced into ES cell lines E14.1 and R1
(Hooper et al., 1987; Nagy et
al., 1993
). Targeted clones
were enriched by positive-negative selection and identified by Southern blot
and PCR analysis. DNA sequencing confirmed the precise homeobox swap. Four out
of 100 R1 clones and 16 out of 300 E14 clones were properly targeted, and one
of each ES cell type was used to make chimeras. No difference in phenotypes
between the two was observed. The Neo marker gene was removed by
mating with transgenic mice expressing NlsCre ubiquitously at an
early embryonic stage (kindly provided by Dr Wojteck Auerbach).
Histology
Tissues were fixed in 4% paraformaldehyde overnight, dehydrated and then
embedded in paraffin. Kidneys were sectioned frontally (5 µm) and stained
using a Periodic Acid Schiff (PAS) kit (Sigma). Testes were stained with
Hematoxylin and Eosin. Female reproductive tracts were sectioned and also
stained with Hematoxylin and Eosin.
Alizarin staining of adult skeletons
Skeletons of 4-week-old animals were prepared and stained as previously
described (Selby, 1987; Small
and Potter, 1993
).
Affymetrix gene chip analysis
Uterus tissue was from the uterus horn above the uterus horn joint junction
and below the uterus-oviduct junction, and cervix tissue was collected from
the region below the uterus corpus and above the vaginal hymen. RNA was
prepared using RNAzol reagent (Tel-test). Preparation of biotinylated RNA,
hybridization, washing, staining and scanning of Affymetrix GeneChip probe
arrays were carried out according to Affymetrix protocols. Data was analyzed
with Affymetrix software.
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RESULTS |
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Hoxd11-/- (D11-/-) mice provided
the most sensitive measure of A11 function. The paralogous
A11 and D11 genes are functionally redundant in the
development of the axial skeleton, limbs, kidneys and reproductive tracts
(Davis et al., 1995). Mice
with A11-/- or D11-/- mutations, for
example, have fairly normal development of the kidney and forelimb zeugopod
(ulna and radius), but in A11-/- D11-/- double
homozygous mutants the kidneys are absent or severely reduced in size, and the
zeugopod of the forelimb is almost entirely missing.
The A11 gene with an A13 homeobox provided near
wild type function in the development of the kidneys, male reproductive tract
and axial skeleton
A11-/- D11-/- double mutant mice commonly
suffer perinatal death from kidney failure (Davis et al.,
1995). The presence of one
wild-type allele (A11+/- D11-/- or
A11-/- D11+/-) restores near normal
kidney development and survival. To define the function of
A1113hd in kidney development, survival rates of
A1113hd/- D11-/- mice were determined
and kidney histology examined. A control cross between double heterozygous
A11+/- D11+/- mice gave 278 progeny,
of which only eight (2.88%) A11-/-
D11-/- pups were found. Predicted additional double
mutants presumably died shortly after birth and were eaten. Only two (0.72%)
A11-/- D11-/- mice survived to
postnatal day (P) 30. By contrast, a cross between
A1113hd/+ D11+/- and
A11+/- D11+/- produced 280 pups of
which 19 (6.79%) were A1113hd/-
D11-/-, with 14 (5%) surviving to P30. This compared well
with the Mendelian predicted percentage of 6.25%. The
A1113hd allele therefore restored near normal survival.
Furthermore, the A1113hd/- D11-/-
kidneys appeared grossly normal, except for the reproducible presence of an
indentation in the anterior region of the left kidney
(Fig. 3A), and with one pair of
kidneys, of eight pairs examined, having small cysts visible on the surface
(data not shown). Histologically the A1113hd/-
D11-/- kidneys were also near wild type, with fewer
dilated distal tubules and more distinct proximal tubule lumens than observed
in A11-/- D11-/- kidneys
(Fig. 3B). However, the medulla
layer of kidney was as severely reduced in size and disorganized in the
A1113hd/- D11-/- mutant as in the
A11-/- D11-/- mutant when compared
with wild type (Fig. 3B).
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The homeobox swapped A1113hd allele also provided
wild-type function in the development of the male reproductive tract.
A11-/- D11+/+ males are generally
sterile with undescended testes and a ductus deferens that is anteriorized to
resemble the epidydimis (Hsieh-Li et al.,
1995).
A1113hd/13hd D11+/+ and
A1113hd/- D11+/- males were fertile,
with descended testes and mature sperm in seminiferous tubules
(Fig. 4A), and their ductus
deferens lacked the tortuosity typical of the anteriorized ductus deferens of
the A11-/- D11+/+ mouse (Hsieh-Li et
al., 1995
;
Fig. 4B).
|
A11-/- D11+/+ mutants show
posteriorization of the 13th thoracic segment into the first lumbar, and
anteriorization of the first sacral segment to a sixth lumbar (Small and
Potter, 1993). Interestingly,
in all six A1113hd/13hd D11+/+ mice
examined the axial skeleton was normal, without anteriorization or
posteriorization (data not shown).
In summary, the A1113hd allele provided apparent wild-type function in the development of the kidney, male reproductive tract and axial skeleton, confirming that the targeted allele produces functional mRNA and protein. These observations are consistent with models predicting that all Hox-encoded proteins bind identical or functionally equivalent downstream targets. However, in examining the limbs and female reproductive tracts, quite different results were obtained.
The A1113hd allele provided antagonizing function
in the development of the limbs
In the developing hindlimb, the A1113hd allele
antagonized normal A11, D11 function. The A11 gene is
expressed in the zeugopod (tibia and fibula in the hindlimb), while the
A13 gene is expressed more distally, in the autopod (paw; Yokouchi et
al., 1991; Haack and Gruss,
1993
; Small and Potter,
1993
). At least five mice were
examined for each of the following genotypes: A1113hd/+
D11+/+, A1113hd/+
D11+/-, A1113hd/+
D11-/-, A1113hd/-
D11+/+, A1113hd/-
D11+/-, A1113hd/-
D11-/- and A1113hd/13hd
D11+/+. The A1113hd allele gave more
severe phenotypes than A11-. The
A11+/- D11+/+ hindlimb was the same as
wild type on the genetic background used in these studies. In the zeugopod
region of the hindlimb, the A1113hd/+
D11+/+ mutants showed a distinct separation of the distal
tibia and fibula, similar to that seen in the A11-/-
D11+/+ mice, and the A1113hd/13hd
D11+/+ hindlimbs showed an even more pronounced separation
(Fig. 5A). In additional allele
combination genotypes, a substitution of the A11- allele
with the A1113hd allele consistently resulted in more
severe separation of the distal tibia and fibula
(Fig. 5B and data not shown).
In the autopod of the hindlimb, the talus and calcaneus bones of
A11-/- D11+/+ mice appeared normal,
whereas for A1113hd/13hd D11+/+ mice
the talus was malformed (not shown) and the calcaneus truncated to the length
of the talus (Fig. 5C). All
allele combinations with at least one A1113hd showed
calcaneus truncation, whereas in the absence of A1113hd
the calcaneus appeared normal in all allele combinations, except for
A11-/- D11-/-, in which the calcaneus
was shortened and also fused with the fibula (data not shown). The penetrance
of the truncated calcaneus phenotype increased with increasing dosage of
A1113hd and to a lesser extent, A11-
and D11- alleles. For example, the truncated calcaneus was
seen in 1/14 (7%) of A1113hd/+
D11+/+, in 4/12 (33%) of
A1113hd/- D11+/+, in 8/11
(72%) of A1113hd/- D11+/- and
in 10/10 (100%) of A1113hd/-
D11-/- mice. All mice with two
A1113hd alleles showed the truncated calcaneus
phenotype.
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The A1113hd allele also had antagonizing effects on
forelimb development. Although the A1113hd/+
D11+/+ forelimb was normal, the
A1113hd/13hd D11+/+ forelimb was more
severely malformed than the A11-/-
D11+/+ forelimb. In the zeugopod region, the ulna and
radius were about one half of normal length, somewhat resembling the three
allele null mutant A11-/- D11+/-,
although with distinctive shapes (Fig.
6A). The styloid apophyses, which were only very mildly affected
in A11-/- D11+/+ mutants were reduced
and/or fused to the ulna and radius in A1113hd/13hd
D11+/+ mice (Fig.
6B, small arrows), approaching the severity of the
A1113hd/- D11-/- or
A11-/- D11-/- mutants
(Fig. 6B, large arrows). In
addition, similar to the hindlimb zeugopod, a substitution of the
A11- allele with the A1113hd allele in
other allele combinations generally resulted in more severely shortened and
malformed forelimb zeugopod (data not shown), with the exception of the
A1113hd/- D11-/- mice, which had
relatively more normal forelimb development than the
A11-/- D11-/- mice
(Fig. 6A). This exception could
reflect some normal limb development function of the
A1113hd allele, which remained hidden in the presence of
wild-type A11 or D11 alleles. Alternatively, the null
A11 mutation, with a deletion and an insertion of a PGK-Neo,
may have produced subtle alterations in the expression patterns of flanking
Hox genes not present with the A1113hd allele during limb
development. It is interesting to note that ectopic expression of
Hoxa13 or Hoxd13 in the chick zeugopod (Yokouchi et al.,
1995; Goff and Tabin,
1997
), or ectopic expression
of Hoxd13 in the mouse zeugopod (van der Hoeven et al.,
1996
; Herault et al.,
1997
; Peichel et al.,
1997
) also results in
shortening of this limb element, similar to the effect observed for
A1113hd.
|
The A1113hd allele assumes A13 function
in development of the female reproductive tract
The A1113hd allele caused a partial homeotic
transformation of the uterus to the more posterior cervix/vagina. The
A11 gene is expressed in the developing uterus and cervix, while
expression of A13 is more posteriorly restricted, to the cervix and
vagina (Taylor et al., 1997;
Post and Innis, 1999
). Mice
examined were 4.5 weeks of age, and estrous cycle matched (Laboratory,
1966
). The lining of the
wild-type uterus consists of a single layer of columnar epithelial cells,
while the lining of the wild-type cervix is many cells thick, making a
squamous epithelium. In contrast to A11-/-
D11+/+ mice, in the A1113hd/13hd
D11+/+ mutants the uterine lining resembled that of the
wild-type cervix (Fig. 7, arrows). This transformation extended throughout most of the uterus, with the
columnar to squamous transition normally present at the uterus-cervix junction
in wild-type mice shifted anterior to near the uterus-oviduct junction in
A1113hd/13hd D11+/+ mutants. The
stromal layer of the mutant uterus also approximated that of the wild-type
cervix, with lower cell density and more fibrous tissue
(Fig. 7, asterisks). The
A1113hd/13hd D11+/+ mutants also
lacked uterine glands and were sterile. While A11+/-
D11+/+ mice are fertile, females with even a single
A1113hd allele were missing uterine glands and sterile. It
was therefore necessary to carry the homeobox swapped A11 gene with
the Neo insertion, which gave a null recessive phenotype, and then to
remove the Neo by germline Cre activity in the last step of
breeding. Because of A1113hd/+ female infertility, it was
extremely difficult to make mice with the A1113hd/13hd
D11-/- genotype.
|
The homeotic transformation of the A1113hd/13hd
D11+/+ uterus was confirmed at the molecular level with
Affymetrix gene chip probe arrays. Murine genome U74A gene chips, with
approximately 12,000 genes, were used to measure gene expression levels in the
4.5-week-old wild-type uterus, wild-type cervix and
A1113hd/13hd D11+/+ uterus. The
resulting molecular fingerprints were then used to determine if the
A1113hd/13hd D11+/+ uterus was shifted
towards the cervix in character. The transcript profile of the mutant uterus
created a detailed molecular portrait showing clear posteriorization. Over 30
genes normally expressed in the cervix but not the uterus were found
transcribed in the A1113hd/13hd D11+/+
uterus. This list contained several keratin genes associated with squamous
epithelium, including K16, K6, K6ß, K14 and, notably,
K13, a marker of the ectocervical epithelium in the female
reproductive tract (Gorodeski et al.,
1990
), and several other genes
of interest (see Appendix). Other genes, normally expressed in the wild-type
uterus and absent in cervix, were inactive in the mutant uterus, again
consistent with posteriorization to cervix. This list included KAP
and calbindin-28, both of which show estrogen responsive expression
in the normal uterus (Meseguer et al.,
1989
; Gill and Christakos,
1995
; Runic et al.,
1996
), and the
decysin gene, which encodes a metalloprotease (Mueller et al.,
1997
). In addition to these
qualitative on/off differences there were many quantitative changes in the
mutant uterus gene expression levels diagnostic of posteriorization
(Appendix). Hoxa13 expression was not detected in either the mutant
uterus or wild-type uterus. In total, comparison of wild-type cervix versus
wild-type uterus gave 106 genes with a tenfold or greater expression level
difference, while comparison of mutant uterus versus wild-type uterus gave 108
genes with over tenfold difference. The two lists of differently expressed
genes shared 54 genes, or about half, consistent with the incomplete homeotic
transformation of mutant uterus to cervix observed at the level of
histology.
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DISCUSSION |
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In the kidney, male reproductive tract and axial skeleton, the
A1113hd allele provided near wild-type function,
consistent with models predicting identical or functionally equivalent
downstream target genes for all Hox encoded proteins. In the developing limbs,
however, the homeobox swapped A11 gene gave antagonizing function.
This could result from a dominant negative effect, with the
A1113hd encoded protein binding to the same downstream
gene targets as for A11/D11, but with opposite effect (e.g.
repression versus activation). Alternatively, the A1113hd
protein could bind to different targets, perhaps those of A13,
leading to distinct developmental outcome. Of interest, according to the
`posterior prevalence' rule in Drosophila, Hox genes located at more
5' positions in the Hox clusters, and expressed in more posterior
domains, are dominant over more 3' genes. This rule predicts that the
more 5' A13 gene, or a homeobox swapped A11 gene with
A13 function, would be dominant over a wild-type A11 gene.
It has been shown that ectopic expression of Hoxa13 or
Hoxd13 (but not Hoxa4) in the zeugopod region in the chick
resulted in zeugopod truncation (Yokouchi et al.,
1995; Goff and Tabin,
1997
). In addition, ectopic
expression of Hoxd13 in the developing zeugopod in mice also resulted
in reduction of this limb element (van der Hoeven et al.,
1996
; Herault et al.,
1997
; Peichel et al.,
1997
), similar to what we
observed in mutants with the swapped A11 gene. This suggests that the
A1113hd allele assumed Hoxa13 function in the
limb and antagonized function of the group 11 Hox genes, probably using the
same mechanism that controls the posterior prevalence phenomenon in normal
development. It is notable that severe zeugopod truncation was observed in the
forelimbs but not in the hindlimbs in our mutants. This could be due to a
quantitative insufficiency of antagonizing activity in the hindlimb, as an
additional group 11 Hox gene, Hoxc11, is normally expressed in the
hindlimb, but not forelimb (Peterson et al.,
1994
; Hostikka and Capecchi,
1998
).
It is interesting to note that possible antagonizing interactions between
the paralogous group 13 and 11 Hox genes were also previously observed in
kidney development. Insertion of a Hoxd9/lacZ construct into the
5' region of the HoxD complex causes ectopic expression of
Hoxd13, resulting in kidney agenesis that resembles the agenesis
found in mice without A11 and D11 function (Kmita et al.,
2000). This result contrasts
with our observation that the A1113hd allele drives near
normal kidney development. The most likely explanation is that the induced
misexpression of Hoxd13 does not properly recapitulate normal Hox
group 11 expression in terms of cell type, timing and expression levels,
therefore perturbing rather than promoting kidney development. It is also
possible that expression of the entire Hoxd13 protein gives different
developmental consequences in the kidney than expression of the A11
protein with a group 13 swapped homeodomain.
The homeotic transformation of uterus towards cervix/vagina clearly
indicated that in this tissue the A1113hd allele assumed
A13 function. The Hox code model of Lewis (Lewis,
1978) predicts that Hox null
mutations will result in anteriorizations and Hox ectopic expression will
drive posteriorizations. Null mutations of A11 and A10 have
been previously reported to anteriorize the uterus towards oviduct (Satokata
et al., 1995
; Gendron et al.,
1997
), and mutation of
A13 anteriorizes the cervix/vagina towards uterus (Post and Innis,
1999
). The
A1113hd allele appeared to effectively give ectopic
expression of A13 function in the uterus, causing it to posteriorize
to cervix/vagina, where the A13 gene is normally expressed. These
results suggest patterning function for Hox genes in the development of the
female reproductive tract.
Distinct segment identity functions have also been defined for Hox genes in
the developing rhombomeres of the mammalian hindbrain. Both misexpression
(Alexandre et al., 1996; Bell
et al., 1999
) and targeted
mutation (Chisaka et al.,
1992
; Carpenter et al.,
1993
; Goddard et al.,
1996
; Studer et al.,
1996
; Gavalas et al.,
1997
; Gavalas et al.,
1998
; Rossel and Capecchi,
1999
) studies support a Hox
code model. For example, ectopic expression of Hoxb1 results in the
homeotic transformation of rhombomere 2 to rhombomere 4 (Bell et al.,
1999
). These results are
consistent with those described in this report, and are again difficult to
reconcile with models predicting that Hox gene function is restricted to the
regulation of cell proliferation.
The genes altered in expression in the mutant uterus are candidate downstream targets of the A13 gene. The single initial difference between wild type and mutant developing uteri was the presence of the A1113hd allele. Expression of this swapped homeobox gene dramatically shifted the gene expression profile of the mutant uterus towards that of the cervix/vagina. The differently expressed genes therefore represent the combination of direct and indirect targets of the A1113hd allele. Moreover, as the identity of the mutant uterus is shifted towards the cervix/vagina, where A13 is normally expressed and as the A1113hd and A13 alleles encode identical homeodomains, these genes are also excellent downstream target candidates for A13 itself. Many of the target genes appear to have functions not related to the regulation of cell proliferation.
It is interesting to note that mutation of Hoxc13, a paralog of
A13, gives a hairless mouse (Godwin and Capecchi,
1998). It has been suggested
that `Hoxc13 could directly control transcription of hair keratin
genes' (Godwin and Capecchi,
1998
). The A13 and
Hoxc13 genes encode homeodomains identical in 55 out of 60 amino
acids. The observed increased expression of a number of keratin genes in the
A1113hd/13hd uterus further indicates that the Hox genes
of the 13 paralogous group can regulate keratin genes.
It has previously been reported that the Hoxa3 and Hoxd3
encoded proteins are functionally equivalent (Greer et al.,
2000). This added to evidence
indicating strong functional redundancy between Hox paralogs (Condie and
Capecchi, 1994
; Davis et al.,
1995
; Horan et al.,
1995
; Fromental-Ramain et al.,
1996a
; Fromental-Ramain et
al., 1996b
). In this report,
however, we show that even the homeoboxes of two contiguous Abd-b
type Hox genes are not functionally interchangeable in all developing
tissues.
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APPENDIX |
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|
Cornified cell envelope components
SPRR1b protein, SPRP 3, Repetin, Loricrin
Transcription factors
Desmosomal adhering junction proteins
Plakophilin 1, Desmocollin 2
Others
Accession numbers
Al604345, Al119347, Al173973, AW259538, AA716963, AA726579, Al118078,
AV233274, Al060798, AW123650, AA794189
Proteins present in wild-type uterus, but absent from wild-type
cervix and A1113hd/13hd D11+/+
uterus
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ACKNOWLEDGMENTS |
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REFERENCES |
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