Download The distribution of E-cadherin during Xenopus laevis development

Development 111. 159-169 (1991)
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159
The distribution of E-cadherin during Xenopus laevis development
GIOVANNI LEVI1, BARRY GUMBINER2 and JEAN PAUL THIERY1
1
Laboratoire du Physiopathologte du Developpement, CNRS URA 1337 and Ecole Normale Supcrtcure, 46 me d'Ulm, 75230 PARIS Cedex,
France
'Department of Pharmacology, University of California, San Francisco, California 94143, USA
Summary
A vast amount of experimental evidence suggests that
cell surface molecules involved in cell-to-cell and/or cellto-substrate interactions participate in the control of
basic events in morphogenesis. E-cadherin is a cell
adhesion molecule directly implicated in the control of
Ca2+-dependent interactions between epithelial cells.
We report here the patterns of expression of E-cadherin
in developmental stages of Xenopus laevis ranging from
early embryo to adult using immunofluorescence microscopy. Although its distribution shares some similarities with those of L-CAM in the chicken and
E-cadherin/Uvomorulin in the mouse, the distribution
of E-cadherin in Xenopus presents several peculiar and
unique features.
In early stages of Xenopus development, E-cadherin is
not expressed. The molecule is first detectable in the
ectoderm of late gastrulas (stage 13-13.5 NF). At this
time both the external and the sensory layer of the nonneural ectoderm accumulate high levels of E-cadherin
while the ectoderm overlying the neural plate and
regions of the involuting marginal zone (IMZ) not yet
internalized by the movements of gastrulation are
E-cadherin-negative. Unlike most other species, endodermal cells express no or very low levels of E-cadherin
up to stage 20 NF. Endodermal cells become strongly
E-cadherin-positive only when a well-differentiated
epithelium forms in the gut. No mesodermal structures
are stained during early development. In the placodes,
in contrast to other species, E-cadherin disappears very
rapidly after placode thickening.
During further embryonic development E-cadherin is
present in the skin, the gut epithelium, the pancreas,
many monostratified epithelia and most glands. Hepatocytes are stained weakly while most other tissues,
including the pronephros, are negative. In the mesonephros, the Wolffian duct and some tubules are
positive.
During metamorphosis a profound restructuring of
the body plan takes place under the control of thyroid
hormones, which involves the degeneration and subsequent regeneration of several tissues such as the skin
and the gut. All newly formed epithelia express high
levels of E-cadherin. Surprisingly, degenerating epithelia of both skin and intestine maintain high levels of
the protein even after starting to become disorganized
and to degenerate.
In the adult, staining is strong in the skin, the glands,
the lungs, the gut epithelium and the pancreas, weak in
the liver and absent from most other tissues.
Our results show that the expression of E-cadherin in
Xenopus is strongly correlated with the appearance of
differentiated epithelia.
Introduction
different levels of the same cadherin can specify the
adhesive properties of a cell, leading to the selective cell
aggregation (Nagafuchi et al. 1987; Edelman etal. 1987;
Hatta et al. 1988; Nose et al. 1988; Mege et al. 1988;
Miyatani et al. 1989; Friedlander et al. 1989; Matsuzaki
et al. 1990). A number of perturbation experiments
(Gallin et al. 1986; Dubande/a/. 1987; Nose etal. 1988;
Detrick et al. 1990) provide strong support for the
hypothesis that cadherins are essential for the control of
critical events of morphogenesis (Edelman, 1985, 1986;
Takeichi, 1988; Thiery, 1989).
So far three cadherins have been identified in
Xenopus laevis. (1) Xenopus E-cadherin was identified
in the Xenopus A6 epithelial cell line by antibody cross-
Cadherins are a family of homophilic Ca2+-dependent
cell adhesion molecules (Takeichi, 1988); they share
partial identity sequence and are highly conserved
through evolution (Gallin etal. 1985, 1987; Nagafuchi et
al. 1987; Nose et al. 1987; Hatta et al. 1988; Miyatani et
al. 1989; Detrick etal. 1990). In the same species several
cadherins coexist (e.g. E-, P-, and N-cadherin in the
mouse) each with a unique tissue distribution and
binding specificity which is determined by restricted
regions of the amino-terminal part of the molecule
(Nose et al. 1990). Transfection experiments show that
the expression of different types of cadherins or of
Key words: Xenopus laevis, cadherins, cell-cell adhesion,
calcium-dependent interaction, immunofluorescence
microscopy.
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G. Levi, B. Gumbiner and J. P. Thiery
reactivity with the homologous molecule in dog.
Several monoclonal antibodies have been prepared
against this molecule but, so far, no sequence data are
available (Choi and Gumbiner, 1989). Xenopus
E-cadherin has a relative molecular mass of 140x10 MT
and shares several biochemical characteristics with
mouse E-cadherin such as the Ca2+-dependent resistance to trypsin. It is expressed primarily in epithelial
tissues (Choi and Gumbiner, 1989). (2) A 140xlO 3 ^,.
protein, highly homologous to mouse and chicken
N-cadherin, is expressed in Xenopus mesoderm and
nervous system shortly after induction (Detrick et al.
1990). The ectopic expression of this molecule in the
ectoderm leads to the formation of cell boundaries and
to severe morphological defects. (3) Finally a
HOxlOi3 MT cadherin-like protein has been recently
identified in Xenopus eggs and cleaving embryos. This
cadherin has been proposed to play a role in the Ca2+dependent adhesion between cleavage stage blastomeres. Its possible relationship to mouse P-cadherin is
not yet known (Choi et al. 1990).
In this paper we have determined the distribution of
E-cadherin at all stages of Xenopus development
paying particular attention to variations of its expression in regions of the embryo where the morphogenetic process is more active.
Materials and methods
Animals
Sexually mature Xenopus laevis were obtained from the
Service d'Elevage de Xenope of the Centre National de la
Recherche Scientifique (Montpellier). Embryos were obtained by artificial fertilization. Embryos were stripped from
females injected 12 h earlier with human chorionic gonadotropin (1000i.u.; Sigma Chemical Co., St. Louis, MO) and
fertilized with the minced testis of males injected 12 h earlier
with 400i.u. of human chorionic gonadotropin. The embryos
were maintained in 10% filter-sterilized Holfreter's solution
at room temperature. Animals at different stages through
metamorphosis were purchased from Nasco (Fort Atkinson,
Wisconsin) and were either used immediately upon delivery
or maintained at 24°C and fed twice weekly with Nasco
Xenopus brittle. Stages of development were determined
according to Nieuwkoop and Faber (1967).
Antibodies
The preparation and the characterization of the monoclonal
antibodies to Xenopus laevis E-cadherin used in this study has
been previously described in detail (Choi and Gumbiner,
1989). Most of the results were obtained using the monoclonal
antibody 5D3 alone; however, no difference in staining
pattern or intensity was detected using a mixture of four
monoclonal antibodies (5D3, 8C2, 19A2 and 31D1) all
directed against Xenopus laevis E-cadherin. In all cases we
made use of diluted ascites fluids.
Immunohistochemistry
Paraffin sections were prepared for staining using a previously
published procedure (Levi et al. 1987; Gurdon et al. 1976);
when this procedure was used to prepare sections of large
metamorphic animals, the method was modified as described
(Levi et al. 1990). Briefly, whole embryos or dissected organs
were frozen in isopentane cooled in liquid nitrogen and
immediately immersed in methanol at —80°C. The samples
were then maintained in methanol at —80°C for periods
ranging from three days .to four months depending on the size
of the animal with weekly changes of cold methanol. The
tissues were serially transferred to methanol equilibrated at
-20°, 4°, and 20°C for at least two hour at each step. The
samples were then immersed twice in xylene until completely
clarified for a total period not greater than 30min and then
transferred to a solution of 50% Paraplast (Monoject
Scientific, St Louis, MO) in xylene at 56°C for 20 min in a
vacuum oven, infiltrated three times with Paraplast at 56°C
under vacuum for 45min, and embedded in Paraplast.
Sections 10 ^m thick were cut using a microtome (Lemardeley, Paris, France), floated on distilled water at 45°C,
collected on washed glass slides and dried on a heating plateC at
45C for at least one hour. To determine the pattern of
distribution of E-cadherin immunoreactivity in embryos
between stage 2 and 20 NF, we collected the complete set of
serial sections corresponding to the whole embryo and stained
them at the same time.
For immunofluorescence, sections were deparaffinized in
Xylene (3x2min), rehydrated and incubated sequentially
with the primary monoclonal antibody (10/igmP 1 in PBS,
5% foetal calf serum (FCS); overnight), biotinylated goat
anti-mouse IgG secondary antibodies (10^gml~' each in
PBS, 5% FCS; 2h) and FITC-conjugated streptavidin
5ugm\ ~] in PBS, 5% FCS; 30 min). In the case of early
embryos, the staining was confirmed by using phycoerythrinconjugated streptavidin (Biomeda Corp., Foster City, CA)
and a barrier filter at 550 nm to reduce the intensity of the yolk
autofluorescence. The sections were observed with a Leitz
epifluorescence microscope. Control sections were stained
either with similar dilutions of monoclonal antibodies directed
against other epitopes or omitting the primary antibody.
Results
Early embryonic development, gastrulation
We could not detect E-cadherin by immunohistofluorescence until stage 12.5. Between stage 12.5 and 13.5
there was a progressive increase in fluorescence in the
non-neural ectoderm, so that by stage 13.5 an intense
signal could be detected in ectodermal cell surface. At
this stage the distribution of the anti-E-cadherin
immunoreactivity was as follows. (1) Staining was
present both in the outer and inner (sensorial) layer of
the ectoderm (Fig. 1 A,C,D,G). All regions of the
ectoderm were stained, with the notable exceptions of
the area of the ectoderm within the developing neural
plate, which was stained very weakly (Fig. 1 B,H), and
a ring of cells surrounding the blastopore, which
possibly corresponded to cells of the involuting marginal zone (prospective mesoderm and endoderm) prior
to invagination (Fig. 1 I). (2) The mesoderm and the
endoderm were not stained. (3) In tangential section
through the ectoderm, E-cadherin immunostaining was
predominantly associated with the cell surface in areas
of cell-cell contact (Fig. 1 D).
Neurulation, larval development
During later development there was a progressive
reduction in the size of the unstained region of the
E-cadherin during Xenopus development
161
lateral pi.
mesoderm
Fig. 1. Distribution of anti-E-cadherin immunoreactivity in Xenopus gastrula. (A-C,G,1) Section corresponding to the
regions designed in the drawings (the left drawing is a sagittal section of the gastrula passing through the neural plate, the
solid line corresponds to the horizontal plane of section which gives rise to the right drawing). (D) Tangential section
through the gastrula ectoderm, (H) Transverse section through the neural plate. (E,F) Phase-contrast images corresponding
to B,C. In stage 13.5-14 Xenopus gastrula E-cadherin is present in most of the ectoderm (A,C,D,G) in the region of
cell-cell contact (D), the ectoderm overlying the neural plate (B.H) and in the region surrounding the yolk plug (I) is not
labeled. The mesoderm and the endoderm are negative. The region of the embryo expressing E-cadherin is indicated by
the chessboard filling in the summary drawings, ar, archenteron; bl, blastocoele; e, ectoderm; en, endoderm; m,
mesoderm; n, notochord; np, neural plate; yp, yolk plug. Magnification: A - C , I , x200; D, X500; G,H, xlOO.
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G. Levi, B. Gumbiner and J. P. Thiery
Fig. 2. Distribution of anti-E-cadherin immunoreactivity in Xenopiis neurula. (A) Stage 18 neurula, the ectoderm overlying
the neural plate region is not stained, B is the phase-contrast image corresponding to A; (C-E) transverse sections at
different levels of a stage 23 embryo; C is the most rostral section and E is the most caudal, a, cement gland; en
endoderm, h, hypophysis; n, notochord; nt, neural tube; s, somite. Magnification: xlOO.
ectoderm within the neural plate so that, by stage 18,
only a small ectodermal region adjacent to the neural
groove was not stained (Fig. 2A). At stage 23 the
epidermis of the entire embryo was strongly E-cadherin
positive (Fig. 2C-E), while the endoderm (Fig. 2D),
mesodermal derivatives (Fig. 2D,E) and the nervous
system were negative. The cement gland and the
hypophysis expressed very high levels of the molecule
from the beginning of their differentiation (Fig. 2C).
The ear placodes develop as a thickening of the
sensorial layer of the ectoderm between stage 21 and
22. Shortly after their formation the level of E-cadherin
expression in the placode epithelium started to diminish
(Fig. 3A). During placode depression (Fig. 3B) and
closure of the ear vesicle (Fig. 3C) all the cells of the
placode, with the exception of those still in contact with
the inner layer of the ectoderm, did not express high
levels of the molecule. After expansion of the ear
vesicle (Fig. 3D), E-cadherin was not detectable in the
auditory organ. A similar rapid down modulation of
E-cadherin expression could be observed during the
formation of the olfactory organ so that invaginating
cells of the nasal pit destined to form the sensory part of
the organ expressed much lower levels of E-cadherin
compared to the surrounding epidermis (Fig. 3E). It is
interesting to note that during placode formation the
invaginating epithelial cells both in the otic and
olfactory placode express high levels of the neural cell
adhesion molecule N-CAM (Levi et al. 1987), which is
not expressed in the epidermis. Therefore a sharp
border is formed between N-CAM- and E-cadherinpositive territories.
Distribution in the skin
The region of the ectoderm corresponding to the
presumptive embryonic epidermis started to express
E-cadherin since stage 13.5 NF. All epithelial cells of
the embryonic, pre-larval and larval skin, including the
outer epithelial layer and the inner sensorial layer of the
embryonic (Fig. 4A) and pre-larval (Fig. 4B) skin and
E-cadherin during Xenopus development
163
Fig. 3. Distribution of anti E-cadherin immunoreactivity during placode formation. Otic placode stage 22 (A), 24 (B), 27
(C) and 37 (D). E-cadherin immunostaining disappears very rapidly from the otic placode after thickening. Olfactory
placode stage 37 (E); E-cadherin immunoreactivity diminishes sharply in the olfactory cells, a, cement gland; olf, olfactory
organ; op, otic placode; ov, otic vesicle; nt, neural tube; ph, pharynx. Magnification: A-C,E, X150; D, x75.
the large unicellular glands of the larval skin (Fig. 4C),
expressed high levels of the molecule. During metamorphosis a complete restructuring of the skin takes place
under the control of thyroid hormones. This process
occurs differently in various areas of the body. In large
areas of the body skin, there is a strong proliferation in
the stratum germinativum which leads to the formation
of a new epidermis beneath the remnants of the larval
epidermis. This new epidermis consists initially of a
monostratified epithelium and of proliferating groups of
cells, which will develop into glands. Other areas of the
skin, such as the tail skin, the skin of the body wall
overlying the developing limbs and several stripes of
skin on the body degenerate, gradually becoming
disorganized and then undergoing a process of cornification. During this whole process, all cells of the
epidermis continued to be strongly immunopositive for
E-cadherin, while no reactivity was detected in the
underlying dermis (Fig. 4D,E,F,H; Fig. 5A-D). Surprisingly, this was true also for areas of degenerating
skin both in the body (Fig. 4G; Fig. 5A-C) and in the
tail (Fig. 5D), even when the epidermis started to be
completely disorganized (Fig. 5B,C). The draining
ducts of both granular and mucus glands in the
postmetamorphic skin were strongly positive while the
glandular epithelium was very weakly stained (Fig. 4
E,F). In the adult skin, strong E-cadherin immunoreactivity persisted in the stratum germinativum of the
epidermis and in the exocrine ducts and diminished
gradually in more external layer of the epidermis
(Fig. 41).
Distribution in the digestive tract and related organs
E-cadherin was not detectable by immunohistofluorescence in any endodermal derivative until a welldifferentiated epithelium had formed. Because the
development of the digestive tract occurs more rapidly
in rostral than in caudal regions, the molecules started
to accumulate at different stages of development in
different portions of the digestive tract. In a stage 41
embryo, for example, the tall columnar epithelium of
the midgut had just formed when E-cadherin staining
appeared as faint, radially oriented lines corresponding
to the cell membranes (Fig. 6A), while the stomach and
the duodenum were already well differentiated and
anti-E-cadherin antibodies stained all epithelial cells
brightly (Fig. 6B). The epithelium of the larval intestine
was strongly labeled by anti-E-cadherin antibodies
(Fig. 6C). This staining persisted through metamorphosis even when' the epithelium started to become
vacuolated (Fig. 6D) and disorganized (Fig. 6E), and
was clearly present also in the reorganized postmetamorphic intestinal epithelium (Fig. 6F).
The monostratified epithelium of the gall bladder as
well as the pancreatic acini and the hepatic, bile and
pancreatic ducts were strongly positive throughout
development while parenchyma! cells of the liver
showed only a faint immunoreactivity. This was most
evident in sections where the relative intensity of
immunoreactivity of these structures could be compared (Fig. 6A,B; Fig. 7A,B). Other positive epithelia
included the gills, developing lungs, the Wolffian duct
and some tubules in the mesonephros; however, the
164
G. Levi, B. Gumbiner and J. P. Thiery
Fig. 4. Distribution of anti-E-cadherin immunoreactivity in the skin. (A) Embryonic skin, stage 19; (B) pre-larval skin,
stage 32; (C) larval skin, stage 51, note the numerous large unicellular glands; (D) premetamorphic skin, stage 55, a new
epidermis has formed beneath the larval skin, and glands are starting to form beneath the epidermis; (E) region of
metamorphosed skin (stage 61) containing mucus glands; (F) granular and mucous glands in postmetamorphic skin (stage
64); (GjH) junction (arrow) between areas of metamorphosing and degenerating skin (G) and metamorphosing and larval
skin (H); (I) adult skin. Magnification: A-E.I x200; G,H, x75.
E-cadherin during Xenopus development
165
Fig. 5. Distribution of anti-E-cadherin immunoreactivity in the limb and tail at metamorphosis. (A,B) Forelimb stage 51,
both the newly forming epidermis of the limb and the degenerating epidermis of the body are E-cadherin positive.
(C) Stage 59. On the left is the metamorphosing skin of a developing forelimb while on the right is a degenerating,
disorganized portion of the body skin; both are labeled by anti-E-cadherin antibodies. (D,E) Degenerating fin tip at stage
59, note the invasion of melanocytes and the cornification of part of the epidermis which continues to be
E-cadherin-positive. lb, limb bud. Magnification: A, x75; C, xlOO; B,D,E, x!50.
pronephros and most tubules of the early mesonephros
were negative.
Distribution in the adult
In adult Xenopus, E-cadherin staining was found in
epithelial tissues lining the surfaces of the body. A very
intense staining was detected in the lungs (Fig. 8A),
skin (Fig. 51), kidney tubules (Fig. 8B), intestinal
epithelium (Fig. 8C) and oviduct epithelium. No
E-cadherin staining was detectable in all other tissues
examined (brain, nerve, skeletal muscle, heart and
aorta). In the adult liver we could detect only a faint
staining on the hepatocytes while the epithelia of the
hepatic and bile ducts were intensely labeled (Fig. 8D).
Discussion
In this study, we examined the distribution of
E-cadherin, a Ca2+-dependent cell adhesion molecule
during the development of Xenopus laevis. Our major
findings are as follows, (a) E-cadherin is not present in
the first stages of Xenopus development (as previously
observed by Choi and Gumbiner, 1989). (b) It is first
detectable by immunohistochemistry in the ectoderm of
stage 13.5 gastrulae only in areas of prospective
epidermal differentiation, (c) During placode development, the intensity of E-cadherin staining diminished
very rapidly after placode thickening, (d) Endodermal
cells express E-cadherin at detectable levels only when
a differentiated columnar epithelium forms in the
digestive tract, (e) During metamorphosis high levels of
166
G. Levi, B. Gumbiner and J. P. Thiery
Fig. 6. Distribution of anti-E-cadherin immunoreactivity in the digestive tract. (A) Midgut of a stage 41 embryo, staining
of the tall columnar epithelium is barely visible; (B) stomach and duodenum of a stage 41 embryo; (C) larval intestine
(stage 53); (D) prometamorphic intestine (stage 59); (E) degenerating intestine during the metamorphic climax (stage 61);
(F) postmetamorphic intestine (stage 64). li, liver; p, pancreas; st, stomach. Magnification: xlOO.
E-cadherin are present in many epithelia; low levels are
present in the liver. E-cadherin persist in the skin and in
the gut epithelia even when they start to become
disorganized and degenerate during the metamorphic
climax, (f) In the adult, E-cadherin is present in the
skin, the gut, the lungs, the glands and the pancreas;
hepatocytes are weakly stained and most other tissues
are negative.
E-cadherin/Uvomorulin was first identified in cleavage stage mouse embryos and plays a critical role in
blastomere compaction in the 8- and 16-cell stage
morulas (Hyafin etal. 1980;Ogouetal. 1983; Damskyet
al. 1983; Shirayoshi et al. 1983; Vestweber and Kemler,
1984; Johnson et al. 1986; Vestweber et al. 1987).
During later development of the mouse embryo, after
being expressed by all cells of the embryo, E-cadherin
disappears from the mesoderm, but persists in the
ectoderm and in most regions of the endoderm
(Damjanov et al. 1986); the molecule is then present in
the liver and most epithelia of the adult (Peyrieras et al.
1983; Vestweber and .Kemler, 1984; Takeichi, 1988).
However, no systematic study of the distribution of
E-cadherin during mouse development has yet appeared. L-CAM is a calcium-dependent cell adhesion
molecule in the chicken (Gallin et al. 1983, 1985, 1987).
It has been repeatedly proposed that L-CAM is the
avian analogue of E-cadherin. However, this possibility
has been recently questioned on the basis of sequence
data (Nose et al. 1990). The distribution of L-CAM
during the embryonic development of the chicken has
been determined in detail: like E-cadherin, L-CAM is
present on most cells of the early embryo and
disappears then from the mesoderm and neural
structures to persist on most limiting epithelia (Thiery
etal. 1984).
Xenopus E-cadherin has been identified on the basis
E-cadherin during Xenopus development
Fig. 7. Distribution of anti-E-cadherin immunoreactivity in
the liver, pancreas and gall bladder. (A) A section through
the liver, bile duct and gall bladder in a stage 53 tadpole;
(B) section through the liver, pancreas and intestine in a
stage 51 tadpole. Although some low-level
immunoreactivity could always be detected in the liver this
was much fainter than that present in the epithelia of the
gall bladder, ducts and pancreas, bd, bile duct; li, liver; gb,
gall bladder; p, pancreas. Magnification x100.
of its cross-reactivity with a polyclonal antibody
directed against mammalian E-cadherin. It shares
several biochemical properties peculiar to mouse
E-cadherin (Choi and Gumbiner, 1989).
The distribution of E-cadherin immunoreactivity in
Xenopus laevis differs from those of E-cadherin in the
mouse and L-CAM in the chicken. First, both mouse-Ecadherin and L-CAM are already present in the very
first stages of embryonic development and have
therefore been called primary CAMs (Edelman,
1988a, b). In Xenopus, E-cadherin appears only in
specific regions of the ectoderm when a well-differentiated epithelium is formed; in the endoderm at
variance with other species no E-cadherin is present
until the gut epithelium starts to form; finally, in several
developing epithelia such as the placodes, the pronephros and the liver, the levels of E-cadherin expression in
Xenopus are much lower than those observed in other
species where E-cadherin is a prevalent molecule.
167
The present data are also different from those
obtained using anti-chicken-L-CAM antibodies on
Xenopus embryos (Levi et al. 1987); beside staining
most epithelia, these antibodies stained diffusely the
early embryo and brightly some structures (such as the
otic vesicle) that are not stained by anti-E-cadherin
antibodies. One possible explanation for the difference
is that anti-L-CAM antibodies might have recognized
more than one cadherin in Xenopus and that their
staining patterns represented the sum of the distributional maps of two or more cadherins. Another
possible explanation for the different distributions
observed is that anti-L-CAM and anti-Xenopus-Ecadherin recognize different cadherins. This would
imply that more than one cadherin coexist on the same
epithelial cell; such a situation occurs in chicken
development where both L-CAM and A-CAM (probably the chicken equivalent of N-cadherin) are occasionally coexpressed by the same cells (e.g. the
endoderm, the otic and lens placode) (Thiery et al.
1982; Duband et al. 1988). Indeed a second Xenopus
cadherin has been identified recently (Choi et al. 1990);
this molecule shares with E-cadherin and all other
known cadherins a common cytoplasmic domain, it is
present in the eggs and cleaving embryos and it has a
relative molecular mass of 120000, the same as that
recognized in Xenopus by anti-L-CAM antibodies (Levi
et al. 1987).
Given the high homology between different cadherins and the possibility that not all cadherins have yet
been described, it is not possible to compare unequivocally the distribution of cadherins in different species
until complete sequence data are available. In this
respect, the situation of Xenopus laevis may be
particularly difficult because it has a pseudotetraploid
genome where polymorphic alleles of the same gene
might have diverged during evolution; this has been
suggested already for Xenopus N-cadherin, which is
present in two closely related variants (Detrick et al.
1990). Furthermore, it has been reported (Detrick et al.
1990) that antibodies raised against fusion proteins
encoding Xenopus N-cadherin recognize several bands
in Western blots of Xenopus brain tissue.
The early distribution of E-cadherin in Xenopus
resembles closely that of a number of epidermal
antigens (Itoh et al. 1988; Jones, 1985). Several studies
suggest that the animal pole region of the embryo at
cleavage stage differentiates autonomously into epidermis (Slack, 1984, 1985; Jones and Woodland, 1986,
1987); however, when the ectoderm is cultured in the
presence of mesoderm, a process of induction leads to
suppression of expression of epidermal markers and
enhances the expression of neural genes such as the
neural cell adhesion molecule N-CAM. Although
expressed in most epidermal regions of the late
gastrula, E-cadherin never accumulated in regions
overlying the neural plate. Therefore, it is conceivable
that, during neural induction, E-cadherin expression is
inhibited in the neural plate by the same set of
molecular signals that activate the expression of
N-CAM.
168
G. Levi, B. Gumbiner and J. P. Thiery
•
4
t*,^ft
9
„.«
B r% W
Fig. 8. Distribution of anti-E-cadherin immunoreactivity in adult organs. (A) Lungs; (B) kidney, the tubules, particularly
surrounding the glomeruli, which were negative, are strongly E-cadherin positive; (C) stomach; (D) liver, the bile duct
epithelium (arrows) is much more strongly stained than the hepatocytes. (g) glomerulus. Magnification: xlOO.
A further point of interest of our data is given by the
persistence of E-cadherin in degenerating epithelia
during metamorphosis. Indeed we could detect a clear
immunoreactivity even when the epithelia started to
become disorganized before falling apart. This is
somehow surprising as it is supposed that E-cadherin is
responsible for the stabilization of cell-to-cell contacts
in the epithelia and one might expect its down
regulation before epithelia disorganization. One must
however, remember that the adhesive function of
E-cadherin can be modulated in several ways; for
example, it is known that in the absence of calcium the
adhesive properties of the molecule are lost, and
several observations indicate that a complex interaction
with cytoskeletal elements is needed to assure cadherin
function (Nagafuchi and Takeichi, 1988; Ozawa et al.
1989; Friedlander et al. 1989; Jaffe et al. 1990); the sole
presence of the protein is not sufficient to assess its
functionality.
This work was in part supported by the Centre National de
la Recherche Scientifique, a grant of the Fondation pour la
Recherche sur les Myopathies (contract AFM, 1990), and by
the National Institute of Health grant No. GM37432. This
work was done under the tenure of an Established Investigatorship from the American Heart Association to Dr Barry
Gumbiner. Dr Giovanni Levi was a recipient of a grant from
the Associazione Italians per la Promozione delle Ricerche
Neurologiche, ARIN.
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(Accepted 25 September 1990)