1 The Jackson Laboratory, 600 Main Street, Bar Harbor, ME 04609, USA
2 Max-Planck Institute of Immunobiology, Stuebeweg 51, D-79108 Freiburg,
Germany
* Author for correspondence (e-mail: bbk{at}jax.org)
Accepted 15 June 2004
![]() |
SUMMARY |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: Mouse, Maternal, E-cadherin, ß-catenin, Embryonic genome, Adhesion, Compaction
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Null mutants of both E-cadherin and ß-catenin exhibit an early
embryonic lethal phenotype. E-cadherin null embryos fail to form an intact
trophectoderm cell layer (Larue et al.,
1994). Lack of ß-catenin results in early gastrulation
lethality, with no mesoderm formation and a block in anterior-posterior axis
formation and head development (Haegel et
al., 1995
; Huelsken et al.,
2000
). It was postulated that these null embryos developed as far
as they did because stable residual maternal proteins or new synthesis from
maternal transcripts could partially rescue the phenotype. However, these
studies did not query the outcome of embryo development in the absence of the
maternal protein.
E-cadherin (uvomorulin), the prototype and founding member of the cadherin
superfamily of calcium-dependent cell adhesion molecules, plays a central role
in cell adhesion and determination of cell shape
(Kemler et al., 1977;
Yagi and Takeichi, 2000
).
E-cadherin has an extracellular domain that allows homophilic interaction with
E-cadherin molecules on neighboring cells. Stable and functional adherens
junctions are formed by interaction of the cytoplasmic tail of E-cadherin with
the catenins, which in turn interact with the actin cytoskeleton
(Nagafuchi, 2001
). E-cadherin
mediates compaction of the individual blastomeres in the 8-cell stage embryo,
and this adhesion triggers the development of the trophectoderm and other
epithelial cell layers at later developmental stages
(Gumbiner, 1996
). In embryos,
mutation of E-cadherin, perturbation of the function of molecules interacting
with E-cadherin, or disturbance of the compaction process, all lead to the
relocation of E-cadherin, and of the molecules interacting with it
(Clayton et al., 1999
;
Ohsugi et al., 1997
;
Pey et al., 1998
).
ß-catenin is one of several intracellular mediators necessary for the
maintenance and function of E-cadherin in cell-cell interactions, as well as
being a central player in the WNT signal transduction pathway
(Kemler et al., 1989). This
dual role is defined by its cellular localization and protein-binding partners
(Gottardi and Gumbiner, 2001
;
Miller and Moon, 1996
).
ß-catenin binds to the cytoplasmic domain of E-cadherin
(Nagafuchi and Takeichi, 1989;
Ozawa et al., 1989
), and to
-catenin, which connects the E-cadherin-catenin adhesion complex with
the actin filament network. ß-catenin is also bound by dynein and may
thus tether microtubules at adherens junctions, ensuring the interactions
between microtubule and actin networks thought to be crucial for mechanical
and signaling events in the cell cortex
(Ligon et al., 2001
).
In somatic cells, ß-catenin is the central component of the
WNT/ß-catenin signal-transduction pathway. Upon WNT receptor-ligand
binding and several intermediate steps, ß-catenin translocates to the
nucleus, and, in association with transcription factors of the TCF/LEF1
family, controls the expression of target genes. In the absence of a WNT
signal cytosolic ß-catenin associates with a multimeric protein complex,
consisting of APC (adenomatous polyposis coli), GSK3B (glycogen synthase
kinase 3ß), CSNK1A1 (casein kinase I) and AXIN, in which it is
phosphorylated and marked for degradation by the ubiquitin-proteasome pathway
(Gottardi and Gumbiner, 2001
;
Liu et al., 2002
).
ß-catenin is also key in a number of intracellular pathways:
ß-catenin binds to transcriptional co-activators and to a component of
the SWI/SNF chromatin-remodeling complex to activate transcription either by
recruiting general transcription factors to target gene promoters, or by
changing chromatin structure (Barker et
al., 2001
; Hecht et al.,
2000
; Miyagishi et al.,
2000
; Nielsen et al.,
2002
; Takemaru and Moon,
2000
).
To determine the role of E-cadherin and ß-catenin during the oocyte to
embryo transition, we used the oocyte specific Zp3-cre transgene
(de Vries et al., 2000), in
combination with proven loxP-tagged (floxed) alleles of these
molecules. The E-cadherin floxed allele has been used to determine the role of
E-cadherin in the lactating mammary gland, the adherens junctions in the
epidermis, hair follicle formation and the peripheral nervous system
(Boussadia et al., 2002
;
Young et al., 2002
;
Young et al., 2003
). The
floxed ß-catenin allele has been successfully used to delineate a role
for ß-catenin in brain development, in development of the ectodermal
ridge and neural crest, in vascular development, and in the embryonic endoderm
(Barrow et al., 2003
;
Brault et al., 2001
;
Cattelino et al., 2003
;
Hari et al., 2002
;
Lickert et al., 2002
;
Machon et al., 2003
). We now
report that deletion of this floxed ß-catenin allele only partially
deletes the ß-catenin gene, resulting in a truncated protein without its
N-terminal part.
Using combinations of the Zp3-cre transgene and floxed alleles, we found that these molecules are crucial for maintaining blastomere adhesion, but that such cell contact is not essential for initiation of development. These results also suggest that the WNT/ß-catenin signaling pathway is probably not functional at this time in development. Interestingly, the absence of maternal E-cadherin in combination with the partially deleted ß-catenin allele results in rescue of the loss-of-embryo phenotype found in females whose oocytes express truncated ß-catenin. A role for E-cadherin-ß-catenin interaction during the oocyte to embryo transition is suggested.
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
Embryo isolation
All embryos were obtained from timed matings of four-week old females, as
described (Evsikov et al.,
2004), ensuring collection of embryos as synchronized in their
development as possible.
Whole-mount immunofluorescence
Whole-mount immunofluorescence was performed as described
(Evsikov et al., 2004). The
primary antibodies were: a monoclonal antibody that recognizes an epitope in
the N-terminal part of ß-catenin (catalog number 610153, BD Transduction
Laboratories); a rabbit polyclonal ß-catenin antibody raised against the
C-terminal part of the protein (Sigma catalog number C-2206); and polyclonal
rabbit antiserum against E-cadherin (GP84)
(Vestweber and Kemler, 1984
).
Secondary antibodies were: CyTM3-conjugated donkey anti-rabbit and
CyTM3-conjugated AffiniPure donkey anti-mouse IgG (Jackson ImmunoResearch
Laboratories, catalog numbers 711-165-152 and 715-165-150, respectively).
Proteasome inhibition
Two-cell embryos from mutant E-cadherin and control females were flushed 24
hours after mating. Some of the embryos were immediately fixed and processed
for whole-mount immunofluorescence. The rest of the embryos were cultured in
vitro in M16 medium, with or without 5 µM N-CBZ-LEU-LEU-AL (MG132; Sigma,
catalog number C-2211; stock solution was 5 mM in DMSO), a membrane permeable
inhibitor of the proteinase activity of the 20S proteasome subunit, at a
concentration known to inhibit polar body extrusion
(Josefsberg et al., 2000;
Mellgren, 1997
). After 10
hours of culture at 37°C, 5% CO2, the embryos were fixed and
processed for immunodetection of ß-catenin using the monoclonal
ß-catenin antibody.
RNA purification and RT-PCR
RNA purification was performed as described
(Oh et al., 2000). For RT-PCR,
RNA was resuspended in water containing 40U rRNasin® (Promega),
and DNA removed using the DNA-freeTM kit (Ambion, catalog number 1906).
Reverse transcriptase reactions were carried out using the SuperScriptTM
Preamplification System (Invitrogen, catalog number 11904-018) according to
the supplier's instructions.
Primer pairs for both ß-catenin and E-cadherin were designed to span an intron-exon boundary:
ß-catenin, 5'-AAGGAAGCTTCCAGACATGC-3'/5'-AGCTTGCTCTCTTGATTGCC-3'; and
E-cadherin, 5'-AAGTGACCGATGATGATGCC-3'/5'-CTTCTCTGTCCATCTCAGCG-3'.
PCR reactions were carried out using aliquots of cDNA containing equal
amounts of RNA, determined by a preceding control PCR on two embryo
equivalents of cDNA using mitochondrial ATP synthase (mt-Atp6)
primers
(5'-TTCCACTATGAGCTGGAGCC-3'/5'-GGTAGCTGTTGGTGGGCTAA-3').
PCR products were usually detected using ethidium bromide staining of agarose
gels. However, to ensure that PCR products were detected when a given
transcript was at its lowest level, Southern hybridization
(Sambrook et al., 1989) was
carried out using E-cadherin or ß-catenin
-[32P]-dCTP
labeled cDNA probes.
To determine whether or not the C-terminal coding sequences of ß-catenin were expressed in embryos lacking the N-terminal part of ß-catenin, a primer pair situated in these sequences was used: 5'GAACAGGGTGCTATTCCACG-3'/5'-GAAAGCCGCTTCTTGTAATCC-3'.
Primer pairs used to determine the presence of transcripts of proteins that interact with ß-catenin were as follows:
All PCR conditions were optimized using the Epicentre FailSafeTM PCR Premix Selection Kit (catalog number FS99060), according to the manufacturer's instructions.
Western blotting
Livers were dissected from 6-week-old male mice that were either
heterozygous for the ß-catenin floxed allele
(ßF/ß) or the ß-catenin deleted allele
(ßF-del/ß), and immediately homogenized in
chilled RIPA buffer [25 mM Tris (pH 8.0), 150 mM NaCl, 0.5% deoxycholate, 0.1%
SDS, 1% IGEPAL, 1 mM EDTA] containing protease inhibitors (Calpain Inhibitor
I, catalog number 1086090; Calpain Inhibitor II, catalog number 1086103;
Bestatin, catalog number 874515; Pefabloc SC Plus, catalog number 1873601;
Roche Applied Science), at concentrations according to the manufacturer's
instructions. One hundred and fifty micrograms of extract was separated on an
8% SDS-PAGE gel, transferred to a HybondTM ECLTM nitrocellulose
membrane (Amersham, Catalog number RPN2020D), and incubated in Tris buffered
saline (TBS, pH 7.6) containing 5% non-fat dried milk (NFDM; Carnation brand)
and 0.1% Tween 20 for 1 hour at room temperature. All subsequent steps were
carried out using TBS containing 5% NFDM and 0.1% Tween 20. The blotted
membrane was incubated with the rabbit polyclonal ß-catenin antibody for
16 hours at 4°C. After multiple washes at room temperature, the membrane
was incubated with a secondary antibody supplied with the ECLTM Western
Blotting Analysis System (Amersham, catalog number RPN2108) for 1 hour at room
temperature. Protein was detected using the ECLTM Western Blotting
Analysis System.
Development of embryos containing a maternal and paternal N-ß-catenin allele
ßF/ßF;cre/Ø
(mutant) and
ßF/ßF;Ø/Ø
(control) females were crossed with
ßF-del/ß;Ø/Ø males. Females and
males were either caged together overnight (natural matings), or for 2 hours
13 hours after injection of human chorion gonadotropin (superovulation).
Embryos were flushed at the 2-cell stage, and cultured to the blastocyst
stage. Experiments with embryos from natural matings were repeated three
times, whereas those from superovulation were only done once.
Statistical methods
Analysis of variance (ANOVA) was used to test the difference among the mean
number of pups per litter from the mutant ß-catenin and control females
([ßF/ßF;cre/Ø],
[ßF/ß;cre/Ø], and
[ßF/ßF;cre/Ø]).
Raw data were used in the ANOVA, as they appeared to meet the assumptions of
normality and homogeneous variances. Where the null hypothesis was rejected,
Tukey's W was used to determine which groups were significantly different from
each other. Significance tests were performed at =0.05. The
2 and proportion tests were done as described
(Devore and Peck, 2001
).
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Live-born pups were obtained from embryos lacking maternal ß-catenin or E-cadherin. To confirm that the intervening sequences between two loxP sites in floxed alleles were removed, PCR analysis was performed on DNA extracted from these progeny. Tail DNA from 122 pups derived from embryos lacking maternal E-cadherin contained the deleted-floxed E-cadherin allele (EF-del) (Fig. 2A), and DNA from 116 pups derived from embryos lacking maternal ß-catenin contained the deleted-floxed ß-catenin allele (ßF-del) (Fig. 2B). These and subsequent data demonstrate complete and effective elimination of floxed sequences by this Zp3-cre transgene.
|
ß-catenin and E-cadherin in blastomere adhesion
The N-terminal part of ß-catenin contains the binding site for
-catenin and some part of the binding site for E-cadherin (armadillo
repeats 3-5). Immunofluorescent analysis of ß-catenin in embryos
expressing truncated ß-catenin, using a monoclonal antibody reacting with
the N-terminal part of the protein, was carried out. Although ß-catenin
was detected at the surface of control blastomeres, the N-terminal part of the
protein is not present at the surface of 2-cell stage blastomeres. Intact
ß-catenin translated from paternal transcripts was first demonstrated at
the 4- to 8-cell stage transition (Fig.
3A).
|
Immunofluorescent detection of E-cadherin in embryos lacking maternally derived protein revealed that there was no detectable E-cadherin at the surface of 2- or 6- to 8-cell stage embryos. The first pinpoints of E-cadherin from the paternal allele were only demonstrable at the morula stage (Fig. 3B). Removing the zona pellucida from these cleavage stage embryos, as in the case of the embryos expressing truncated ß-catenin, resulted in immediate dissociation of individual blastomeres of embryos up to the 8-cell stage. Adhesion of the maternal E-cadherin-deficient embryos did not occur until the morula stage, coinciding with the first detectable protein (Fig. 3B). The levels of fluorescence indicated that less E-cadherin was present in morulae that lacked maternal E-cadherin. Nonetheless, compaction occurred in these embryos, albeit an entire cell division later than in normal embryos.
ß-catenin-E-cadherin interaction and effects on protein localization
E-cadherin localization in embryos expressing truncated ß-catenin
To determine whether absence of intact ß-catenin influenced
localization of E-cadherin on the blastomere surface, 2-, 4- and 8-cell stage
embryos expressing truncated ß-catenin were examined using E-cadherin
antibody (Fig. 4A). E-cadherin
is immunodetected at the surface of 2- and 4-cell stage embryos expressing
truncated ß-catenin, although the zone of detection is not as sharply
demarcated as in wild-type embryos, and adhesion does not occur.
Interestingly, E-cadherin could also be detected in the cytoplasm of 2- and
4-cell stage embryos expressing truncated ß-catenin. In 8-cell stage
embryos containing truncated and intact ß-catenin, E-cadherin is
detectable at the cell surface in a normal pattern
(Fig. 4A).
|
To determine whether the truncated ß-catenin protein is also able to translocate to the nucleus, we made use of embryos from EF/EF;ßF/ßF;cre/Ø females. These embryos, which lack maternal E-cadherin and express truncated ß-catenin, were analyzed by immunofluorescence using the polyclonal antibody to ß-catenin. This showed that truncated ß-catenin is found in the pronuclei and nuclei of zygotes and 2-cell stage embryos (Fig. 4C). This result was confirmed using another polyclonal antibody (AbCam, catalog number ab6302), which recognizes the C-terminal part of the protein. Comparison of the pattern of staining between embryos lacking maternal E-cadherin and expressing wild-type ß-catenin, and that of embryos lacking both maternal E-cadherin and expressing truncated ß-catenin reveals a similar staining pattern.
Regulation of ß-catenin levels in embryonic blastomeres
Degradation of ß-catenin via the proteasome in pre-implantation embryos
To determine whether ß-catenin concentration in early embryos could be
controlled by the same mechanism as in somatic cells, we made use of embryos
lacking maternal E-cadherin. As the somatic mechanism involves the proteasome,
2-cell stage embryos were incubated with the proteasome inhibitor MG132 and
ß-catenin was detected using the monoclonal ß-catenin antibody
(Fig. 5A). ß-catenin is
visible at the cell surface and in the cytoplasm of a control embryo
(Fig. 5A, first panel).
ß-catenin protein is detectable in the cytoplasm and especially the
nucleus, but not at the surface of the blastomeres, of an embryo lacking
maternal E-cadherin that was incubated in the proteasome inhibitor
(Fig. 5A, second panel). Low
levels of ß-catenin, slightly more than background staining, are visible
in the nucleus and cytoplasm of an MG132-free, control embryo lacking maternal
E-cadherin (Fig. 5A, compare
panels three and four).
|
Expression of truncated ß-catenin influences embryo development
To determine whether embryo development was adversely affected by the
expression of a maternal and paternal N-ß-catenin
allele, we crossed
ßF/ßF;cre/Ø
females with ßF-del/ß;Ø/Ø males,
giving rise to embryos of the genotype
ßF-del/ßF-del and
ßF-del/ß. As controls,
ßF/ßF;Ø/Ø
females were crossed with
ßF-del/ß;Ø/Ø males. Embryo
development to the blastocyst stage was monitored
(Table 1). Using the
chi-squared (2) test, we determined that embryos obtained from
ßF/ßF;cre/Ø
females developed less efficiently than those from control females.
Hypothetically, if the expression of two deleted-floxed ß-catenin alleles
is lethal, at least 50% of embryos should die. This hypothesis was rejected
using a proportion test, and therefore the loss of embryos from
ßF/ßF;cre/Ø
females cannot be ascribed to the expression of two deleted-floxed
ß-catenin alleles. These data suggest that full-length ß-catenin is
not a requirement for pre-implantation development to blastocyst in vitro.
|
|
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
Although maternal E-cadherin was previously found to be present in the
E-cadherin null embryos, the membrane localization of -catenin,
ß-catenin and ZO-1 (TJP1 Mouse Genome Informatics), all of which
are involved in interacting with E-cadherin to establish proper cell-cell
interaction and organize the cortical actin filament, was abnormal
(Ohsugi et al., 1997
).
E-cadherin null embryos, while containing maternal E-cadherin, are not able to
maintain compaction and form an intact trophectoderm layer
(Larue et al., 1994
). Embryos
lacking maternal E-cadherin undergo compaction and subsequent cavitation as
soon as the paternal protein is expressed. E-cadherin is thus not required for
early embryo development, but synthesis and post-translational modification of
E-cadherin, translated from the newly activated embryonic genome, as well as
its interaction with ß-catenin, is required for the dynamic changes
necessary for blastomere-blastomere adhesion during compaction and
trophectoderm formation.
ß-catenin and pre-implantation embryo development
The floxed ß-catenin allele we used in this study has been used to
delineate the role of ß-catenin and the WNT pathway in development of the
brain, neural crest and embryonic endoderm
(Brault et al., 2001;
Hari et al., 2002
;
Lickert et al., 2002
;
Machon et al., 2003
). Our
analysis revealed that excision of the floxed sequences removes the N-terminal
part of the protein, leaving the C-terminal part intact. The N-terminal part
contains binding sites for
-catenin, GSK3ß, CSNK1A1, Reptin52 and
RUVBL1 (Pontin52). From the best estimates in the literature, the binding
sites for E-cadherin, TCF/LEF, APC and AXIN are partially removed, but in the
cases cited above the WNT/ß-catenin signaling pathway is non-functional.
Because oocytes expressing truncated ß-catenin undergo normal maturation,
fertilization and early 2-cell stage development, we suggest that the
WNT/ß-catenin signaling pathway is not functional at this time in
development. Further support for this is gained from the fact that
ß-catenin translocates to the nucleus in embryos lacking maternal
E-cadherin, in essence mimicking an overexpression of ß-catenin, which in
systems responsive to the WNT/ß-catenin pathway causes detrimental
effects. Nonetheless, these embryos develop normally.
However, it cannot be ignored that the C-terminal part of the protein,
containing binding sites for CREBBP/p300, CATNBP1 (ICAT), SMARCA4 (BRG1) and
SDCCAG33, is still intact. In the nucleus, transcriptional co-activators
p300/CREBBP may bind ß-catenin to activate transcription either by
recruiting general transcription factors to target gene promoters, or by
changing chromatin structure (Hecht et
al., 2000; Miyagishi et al.,
2000
; Takemaru and Moon,
2000
). ß-catenin can also recruit the chromatin-remodeling
factor BRG1 to TCF-responsive promoters, forming a complex to remodel
chromatin and facilitate transcriptional activation
(Barker et al., 2001
;
Nielsen et al., 2002
).
Although this truncated protein might not be active, it is reasonable to
speculate that the C-terminal part of ß-catenin is needed for the initial
changes in chromatin restructuring during nuclear reprogramming that take
place during the oocyte to embryo transition. Truncated ß-catenin, like
the wild-type protein, translocates to the pronuclei of zygotes and the nuclei
of 2-cell stage embryos (Fig.
7C). This may indicate that the truncated protein is able to
interact with different factors, for which transcripts have been shown to be
present in oocytes and early embryos, during this time of nuclear
reprogramming, a hypothesis that remains to be tested.
Females with oocytes that express truncated ß-catenin produce fewer
pups per litter than controls, but this loss-of-embryo phenotype is rescued if
oocytes also do not express maternal E-cadherin
(Fig. 7D,E). It is known that
ß-catenin forms a complex with E-cadherin soon after being synthesized in
the endoplasmic reticulum, and some ß-catenin is left in the free
cytoplasmic form (McCrea and Gumbiner,
1991; Ozawa and Kemler,
1992
). We postulate that in the situation where intact
ß-catenin is newly synthesized from the paternal allele at the late
2-cell embryo stage, it is preferentially sequestered by E-cadherin, resulting
in insufficient amounts of free ß-catenin available for nuclear
translocation. However, in embryos lacking maternal E-cadherin and expressing
the truncated ß-catenin allele, newly synthesized ß-catenin is not
sequestered by E-cadherin, which is absent from the oocyte and which is
activated only at the late 4-cell stage. Intact ß-catenin is thus
available in the free form, and can be channeled to the nucleus
(Fig. 7E).
These results give an intriguing glimpse into the interplay between maternal and newly synthesized E-cadherin and ß-catenin during the oocyte to embryo transition. Early cleavage embryos can therefore be seen as a system that is both robust and able to tolerate quite large changes, while at the same time being dependent on the meticulous timing of new transcription and translation.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Barker, N., Hurlstone, A., Musisi, H., Miles, A., Bienz, M. and
Clevers, H. (2001). The chromatin remodelling factor Brg-1
interacts with beta-catenin to promote target gene activation. EMBO
J. 20,4935
-4943.
Barrow, J. R., Thomas, K. R., Boussadia-Zahui, O., Moore, R.,
Kemler, R., Capecchi, M. R. and McMahon, A. P. (2003).
Ectodermal Wnt3/beta-catenin signaling is required for the establishment and
maintenance of the apical ectodermal ridge. Genes Dev.
17,394
-409.
Boussadia, O., Kutsch, S., Hierholzer, A., Delmas, V. and Kemler, R. (2002). E-cadherin is a survival factor for the lactating mouse mammary gland. Mech. Dev. 115, 53-62.[CrossRef][Medline]
Brault, V., Moore, R., Kutsch, S., Ishibashi, M., Rowitch, D.
H., McMahon, A. P., Sommer, L., Boussadia, O. and Kemler, R.
(2001). Inactivation of the beta-catenin gene by
Wnt1-Cre-mediated deletion results in dramatic brain malformation and failure
of craniofacial development. Development
128,1253
-1264.
Cattelino, A., Liebner, S., Gallini, R., Zanetti, A., Balconi,
G., Corsi, A., Bianco, P., Wolburg, H., Moore, R., Oreda, B. et al.
(2003). The conditional inactivation of the beta-catenin gene in
endothelial cells causes a defective vascular pattern and increased vascular
fragility. J. Cell Biol.
162,1111
-1122.
Clayton, L., Hall, A. and Johnson, M. H. (1999). A role for Rho-like GTPases in the polarisation of mouse eight-cell blastomeres. Dev. Biol. 205,322 -331.[CrossRef][Medline]
de Vries, W. N., Binns, L. T., Fancher, K. S., Dean, J., Moore, R., Kemler, R. and Knowles, B. B. (2000). Expression of Cre recombinase in mouse oocytes: a means to study maternal effect genes. Genesis 26,110 -112.[CrossRef][Medline]
Devore, J. and Peck, R. (2001). Statistics. The exploration and analysis of data. Duxbury: Thomson Learning.
Evsikov, A. V., de Vries, W. N., Peaston, A., Fancher, K., Chen, F., Radford, E., Latham, K., Blake, J., Bult, C., Solter, D. et al. (2004). Systems biology of the 2-cell embryo. Cytogenet. Genome Res. 105,240 -250.[Medline]
Fleming, T. P. and Johnson, M. H. (1988). From egg to epithelium. Annu. Rev. Cell Biol. 4, 459-485.[Medline]
Gottardi, C. J. and Gumbiner, B. M. (2001). Adhesion signaling: how beta-catenin interacts with its partners. Curr. Biol. 11,R792 -R794.[CrossRef][Medline]
Gumbiner, B. M. (1996). Cell adhesion: the molecular basis of tissue architecture and morphogenesis. Cell 84,345 -357.[Medline]
Haegel, H., Larue, L., Ohsugi, M., Fedorov, L., Herrenknecht, K.
and Kemler, R. (1995). Lack of beta-catenin affects
mouse development at gastrulation. Development
121,3529
-3537.
Hari, L., Brault, V., Kleber, M., Lee, H. Y., Ille, F.,
Leimeroth, R., Paratore, C., Suter, U., Kemler, R. and Sommer, L.
(2002). Lineage-specific requirements of beta-catenin in neural
crest development. J. Cell Biol.
159,867
-880.
Hecht, A., Vleminckx, K., Stemmler, M. P., van Roy, F. and
Kemler, R. (2000). The p300/CBP acetyltransferases function
as transcriptional coactivators of beta-catenin in vertebrates.
EMBO J. 19,1839
-1850.
Huarte, J., Belin, D., Vassalli, A., Strickland, S. and Vassalli, J. D. (1987). Meiotic maturation of mouse oocytes triggers the translation and polyadenylation of dormant tissue-type plasminogen activator mRNA. Genes Dev. 1,1201 -1211.[Abstract]
Huelsken, J., Vogel, R., Brinkmann, V., Erdmann, B., Birchmeier,
C. and Birchmeier, W. (2000). Requirement for
beta-catenin in anterior-posterior axis formation in mice. J. Cell
Biol. 148,567
-578.
Johnson, M. H. (1996). Origins of pluriblast and trophoblast in the eutherian conceptus. Reprod. Fertil. Dev. 8,699 -709.[Medline]
Josefsberg, L. B., Galiani, D., Dantes, A., Amsterdam, A. and
Dekel, N. (2000). The proteasome is involved in the first
metaphase-to-anaphase transition of meiosis in rat oocytes. Biol.
Reprod. 62,1270
-1277.
Kanzler, B., Haas-Assenbaum, A., Haas, I., Morawiec, L., Huber, E. and Boehm, T. (2003). Morpholino oligonucleotide-triggered knockdown reveals a role for maternal E-cadherin during early mouse development. Mech. Dev. 120,1423 -1432.[CrossRef][Medline]
Kemler, R., Babinet, C., Eisen, H. and Jacob, F. (1977). Surface antigen in early differentiation. Proc. Natl. Acad. Sci. USA 74,4449 -4452.[Abstract]
Kemler, R., Ozawa, M. and Ringwald, M. (1989). Calcium-dependent cell adhesion molecules. Curr. Opin. Cell Biol. 1,892 -897.[Medline]
Larue, L., Ohsugi, M., Hirchenhain, J. and Kemler, R. (1994). E-cadherin null mutant embryos fail to form a trophectoderm epithelium. Proc. Natl. Acad. Sci. USA 91,8263 -8267.[Abstract]
Lickert, H., Kutsch, S., Kanzler, B., Tamai, Y., Taketo, M. M. and Kemler, R. (2002). Formation of multiple hearts in mice following deletion of beta-catenin in the embryonic endoderm. Dev. Cell 3,171 -181.[Medline]
Ligon, L. A., Karki, S., Tokito, M. and Holzbaur, E. L. (2001). Dynein binds to beta-catenin and may tether microtubules at adherens junctions. Nat. Cell Biol. 3, 913-917.[CrossRef][Medline]
Liu, C., Li, Y., Semenov, M., Han, C., Baeg, G. H., Tan, Y., Zhang, Z., Lin, X. and He, X. (2002). Control of beta-catenin phosphorylation/degradation by a dual-kinase mechanism. Cell 108,837 -847.[Medline]
Machon, O., van den Bout, C. J., Backman, M., Kemler, R. and Krauss, S. (2003). Role of beta-catenin in the developing cortical and hippocampal neuroepithelium. Neuroscience 122,129 -143.[CrossRef][Medline]
McCrea, P. D. and Gumbiner, B. M. (1991).
Purification of a 92-kDa cytoplasmic protein tightly associated with the
cell-cell adhesion molecule E-cadherin (uvomorulin). Characterization and
extractability of the protein complex from the cell cytostructure.
J. Biol. Chem. 266,4514
-4520.
McNeill, H. (2000). Sticking together and sorting things out: adhesion as a force in development. Nat. Rev. Genet. 1,100 -108.[CrossRef][Medline]
Mellgren, R. L. (1997). Specificities of cell
permeant peptidyl inhibitors for the proteinase activities of mu-calpain and
the 20 S proteasome. J. Biol. Chem.
272,29899
-29903.
Miller, J. R. and Moon, R. T. (1996). Signal transduction through beta-catenin and specification of cell fate during embryogenesis. Genes Dev. 10,2527 -2539.[CrossRef][Medline]
Miyagishi, M., Fujii, R., Hatta, M., Yoshida, E., Araya, N.,
Nagafuchi, A., Ishihara, S., Nakajima, T. and Fukamizu, A.
(2000). Regulation of Lef-mediated transcription and
p53-dependent pathway by associating beta-catenin with CBP/p300. J.
Biol. Chem. 275,35170
-35175.
Nagafuchi, A. (2001). Molecular architecture of adherens junctions. Curr. Opin. Cell Biol. 13,600 -603.[CrossRef][Medline]
Nagafuchi, A. and Takeichi, M. (1989). Transmembrane control of cadherin-mediated cell adhesion: a 94 kDa protein functionally associated with a specific region of the cytoplasmic domain of E-cadherin. Cell Regul. 1, 37-44.[Medline]
Nielsen, A. L., Sanchez, C., Ichinose, H., Cervino, M., Lerouge,
T., Chambon, P. and Losson, R. (2002). Selective
interaction between the chromatin-remodeling factor BRG1 and the
heterochromatin-associated protein HP1alpha. EMBO J.
21,5797
-5806.
Oh, B., Hwang, S., McLaughlin, J., Solter, D. and Knowles, B.
B. (2000). Timely translation during the mouse
oocyte-to-embryo transition. Development
127,3795
-3803.
Ohsugi, M., Hwang, S. Y., Butz, S., Knowles, B. B., Solter, D. and Kemler, R. (1996). Expression and cell membrane localization of catenins during mouse preimplantation development. Dev. Dyn. 206,391 -402.[CrossRef][Medline]
Ohsugi, M., Larue, L., Schwarz, H. and Kemler, R. (1997). Cell-junctional and cytoskeletal organization in mouse blastocysts lacking E-cadherin. Dev. Biol. 185,261 -271.[CrossRef][Medline]
Ozawa, M., Baribault, H. and Kemler, R. (1989). The cytoplasmic domain of the cell adhesion molecule uvomorulin associates with three independent proteins structurally related in different species. EMBO J. 8,1711 -1717.[Abstract]
Ozawa, M. and Kemler, R. (1992). Molecular organization of the uvomorulin-catenin complex. J. Cell Biol. 116,989 -996.[Abstract]
Pey, R., Vial, C., Schatten, G. and Hafner, M.
(1998). Increase of intracellular Ca2+ and relocation of
E-cadherin during experimental decompaction of mouse embryos. Proc.
Natl. Acad. Sci. USA 95,12977
-12982.
Sambrook, J., Fritsch, E. and Maniatis, T. (1989). Molecular Cloning: A Laboratory Manual. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press.
Takemaru, K. I. and Moon, R. T. (2000). The
transcriptional coactivator CBP interacts with beta-catenin to activate gene
expression. J. Cell Biol.
149,249
-254.
Vestweber, D., Gossler, A., Boller, K. and Kemler, R. (1987). Expression and distribution of cell adhesion molecule uvomorulin in mouse preimplantation embryos. Dev. Biol. 124,451 -456.[Medline]
Vestweber, D. and Kemler, R. (1984). Rabbit antiserum against a purified surface glycoprotein decompacts mouse preimplantation embryos and reacts with specific adult tissues. Exp. Cell Res. 152,169 -178.[Medline]
Yagi, T. and Takeichi, M. (2000). Cadherin
superfamily genes: functions, genomic organization, and neurologic diversity.
Genes Dev. 14,1169
-1180.
Young, P., Boussadia, O., Berger, P., Leone, D. P., Charnay, P., Kemler, R. and Suter, U. (2002). E-cadherin is required for the correct formation of autotypic adherens junctions of the outer mesaxon but not for the integrity of myelinated fibers of peripheral nerves. Mol. Cell Neurosci. 21,341 -351.[CrossRef][Medline]
Young, P., Boussadia, O., Halfter, H., Grose, R., Berger, P.,
Leone, D. P., Robenek, H., Charnay, P., Kemler, R. and Suter, U.
(2003). E-cadherin controls adherens junctions in the epidermis
and the renewal of hair follicles. EMBO J.
22,5723
-5733.