European Molecular Biology Laboratory, Meyerhofstr 1, 69117 Heidelberg, Germany
* Author for correspondence (e-mail: cohen{at}embl-heidelberg.de)
Accepted 31 October 2002
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SUMMARY |
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Key words: Compartment boundary, Cell affinities, Cell interactions, fringe
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INTRODUCTION |
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The onset of apterous expression in the early wing primordium
induces expression of the Notch ligand Serrate in D cells and restricts
expression of Delta, another Notch ligand, to V cells
(Diaz-Benjumea and Cohen,
1995; Milán and Cohen,
2000
). Dorsally expressed Serrate and ventrally expressed Delta
activate Notch symmetrically in cells on both sides of the DV compartment
boundary (Fig. 1A) (Diaz-Benjumea and Cohen,
1995
; de Celis et al.,
1996
; Doherty et al.,
1996
). Expression of the glycosyltransferase Fringe makes D cells
more sensitive to Delta and less sensitive to Serrate
(Fleming et al., 1997
;
Panin et al., 1997
;
Brückner et al., 2000
;
Moloney et al., 2000
;
Munro and Freeman, 2000
).
Notch activation induces Wg expression in cells along the DV boundary. Later
in development, an increase in dLMO (BX FlyBase) levels reduces Ap
activity in the wing primordium
(Milán and Cohen,
2000
). At this stage, another set of cell interactions takes over
to maintain Wg expression along the DV boundary
(Fig. 1B) (de Celis and Bray, 1997
;
Micchelli et al., 1997
). Wg
induces expression of Serrate and Delta in nearby D and V cells. Serrate and
Delta signal back to activate Notch and thereby maintain Cut and Wg expression
along the DV boundary. The persistent low level of Fringe in D cells continues
to make D cells more sensitive to Delta and less sensitive to Serrate at this
later stage (Milán and Cohen,
2000
).
|
At the time compartments were discovered, it was proposed that lineage
restriction along the compartment boundaries depended on compartment specific
expression of adhesion molecules that conferred differential cell affinities
(García-Bellido et al.,
1973). More recent studies have also indicated a role for cell
communication at the compartment boundary. Engrailed induces expression of the
secreted signaling protein Hedgehog in P cells. Hedgehog acts through Patched
and Smoothened to control gene expression in A cells. Hedgehog signaling is
needed to maintain segregation of A and P compartments
(Blair and Ralston, 1997
;
Rodriguez and Basler, 1997
)
can fulfill this function because it is intrinsically asymmetric. Thus, it is
easy to understand how Hh signaling can induce a difference in cell behavior
at the AP boundary. The situation at the DV boundary is more complex. The role
of Notch signaling is less easy to reconcile with compartment-specific cell
segregation, because Notch is activated symmetrically on both sides of the DV
boundary. Fringe-dependent Notch signaling has been shown to play a role in
segregation of D and V cells (Micchelli
and Blair, 1999
; Rauskolb et
al., 1999
). However, we have previously reported that restoring
Notch activation along the DV compartment boundary is not sufficient to
support the DV boundary under conditions of reduced Apterous activity
(Milán and Cohen,
1999a
). This indicates the need for an additional
Apterous-dependent process that keeps D and V cells apart. The LRR
transmembrane proteins Capricious and Tartan are transiently expressed in D
cells under Apterous control, at which time they contribute to formation of
the boundary between D and V cells (Fig.
1A) (Milán et al.,
2001
). Their function at the DV boundary is transient and they are
subsequently redeployed to produce a difference in affinities between medial
and lateral cells (Milán et al.,
2002
). Thus, maintenance of the DV affinity boundary is
independent of Capricious and Tartan. Notch signaling may contribute to this
process.
In this report we re-evaluate the roles of Apterous and Notch activation in
the DV boundary. Two models have been proposed to explain the roles of Fringe
and Notch. According to one view, dorsal expression of Fringe may have
Notch-independent functions in the generation of an affinity difference
between D and V cells (O'Keefe and Thomas,
2001; Rauskolb et al.,
1999
). Our findings do not support this view and indicate that the
activities of Fringe are mediated through Notch. The second model proposes
that Notch activity induces an adhesive state that is qualitatively modulated
by Apterous to generate dorsal and ventral boundary states
(Blair, 2001
;
Micchelli and Blair, 1999
).
This model is based on the proposal that there is a difference in affinity
between boundary cells and cells within each compartment (wing blade cells).
Our findings do not support this feature of the model. We show that there is
no intrinsic affinity difference between boundary cells and wing blade cells
within a compartment. Only interactions between D and V cells induce an
affinity difference. Instead, we propose that Notch activity cooperates with
Apterous to produce an affinity difference between D and V cells. We present a
model in which the role of Apterous is instructive and the role of Notch is
essential, but permissive.
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MATERIALS AND METHODS |
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Antibodies
Guinea-pig anti-Ap and rat anti-dLMO
(Weihe et al., 2001) were
used. Other antibodies are commercially available.
Genotypes of larvae used for genetic mosaic analysis
Actin>CD2>Gal4; aprk568 females were crossed to
the following males:
To generate clones lacking Ap activity, we made clones lacking the essential co-factor Chip or clones expressing the Ap inhibitor dLMO. Such clones are phenotypically equivalent to removing the ap gene, but can be generated in larger numbers by use of the FRT and flip-out systems. ap is located proximal to the FRT on 2R precluding se of the FRT system to generate large numbers of ap mutant clones.
Measurements of clone shapes
Using NIH Image version 1.60, the perimeter (L) and area (A) of the clones
were measured. The ratio 4 A/L2 was used as a measure of the
shape of the clones. 4
A/L2=1.0 for a perfect circle. Lower
values indicate more irregular shapes. For presentation, 4
A/L2
numbers were rounded off to one significant digit. t-test analysis
was carried out to analyze if the shape of mutant or expressing clones
differed significantly from control clones.
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RESULTS |
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To ask whether the differences between these experiments can be explained in terms of the amount of fringe activity relative to the degree of reduction of ap activity, we compared the ability of different amounts of Fringe expression to rescue three different apterous mutant combinations. apGal4 homozygous larvae show a relatively weak phenotype in wing discs (Fig. 2A). Providing a lower level of Fringe activity using EP3082 did not fully rescue the apGal4 homozygous disc phenotype, whereas providing a higher level of Fringe activity using UAS-fng rescued well (Fig. 2B,C). In the apGal4/aprk568 combination, with intermediate levels of Ap activity, providing the lower level of Fringe activity using EP3082 did not rescue the boundary phenotype. Providing the higher level of Fringe activity using UAS-fng rescued quite well, but not perfectly (Fig. 2D-F). The apGal4/apUG035 combination provided the strongest mutant phenotype. Neither of the Fringe-expressing genotypes rescued the boundary defects, though the UAS-fng was better than EP3082 (Fig. 2G-I). We note that the level of Fringe in both cases was sufficient to generate robust Notch activation, as indicated by Wg expression. As a control, we verified that expression of UAS-Apterous under apGal4 control was sufficient to restore both Notch activation and the DV lineage restriction boundary in the strongest mutant combination (Fig. 2J). Thus, we reiterate our conclusion that Fringe activity in D cells does not appear to be sufficient to generate a DV affinity boundary under conditions of reduced Apterous activity.
Different effects of ubiquitous expression of Apterous or Fringe on
the DV boundary
To further evaluate the contributions of Apterous and Fringe to maintenance
of the DV affinity boundary, we expressed these proteins throughout the P
compartment of the wing disc using engrailed-Gal4. Ectopic expression in P
cells eliminates the difference between D and V cells in the P compartment and
permits an assessment of their effects on the endogenous DV boundary
(visualized by the expression of an ap-lacZ reporter gene). When
UAS-fng was expressed in P cells, Wg expression was lost at the
endogenous boundary between D and V cells in the P compartment, and an ectopic
stripe of Wg expression was induced in cells along the edge of the
engrailed-Gal4 domain (Fig.
3G-I). The interface between D and V compartment cells became
irregular under these conditions. These observations confirm the report by
Rauskolb et al. (Rauskolb et al.,
1999) that a boundary between Fringe-expressing and non-expressing
cells is needed to maintain the DV affinity boundary.
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The effects of Apterous expression differed considerably from Fringe in terms of the relative position of the AP and DV compartment boundaries. In wild-type wing discs, the AP and DV boundaries are perpendicular to each other (Fig. 3A-C). This was also the case when Fringe was expressed in P cells (Fig. 3G-I). The engrailed-Gal4 domain was perpendicular to the DV boundary (in the A compartment), despite expression of Fringe in all P cells. By contrast, expression of Apterous in all P cells caused P cells of ventral origin to relocate into the dorsoposterior quadrant of the disc (Fig. 3D-F). Although these cells were of V compartment origin, they appear to have sorted out into the D compartment by virtue of Ap expression. Under these conditions, the AP and DV compartment boundaries were no longer perpendicular (Fig. 3F). This suggests that Apterous and Fringe do not have comparable abilities to confer D compartment-specific cell behavior. Sorting out can be caused by differences in cell affinity.
Fringe is not comparable with Apterous in its ability to confer D
cell affinity
In genetic mosaics, differences in cell affinity can be visualized by the
shapes of mutant clones (Lawrence et al.,
1999; Liu et al.,
2000
). To compare the effects of Apterous and Fringe on D
compartment cell affinity we examined the shapes of mutant clones lacking
these activities in the D compartment or clones expressing Apterous or Fringe
in the V compartment. Wild-type clones were elongated along the proximodistal
axis of the wing and their borders were irregular
(Fig. 4A), except when they
touched the DV or AP compartment boundaries (see
Fig. 6D). For convenience,
clones of cells lacking Apterous activity were produced either by removing
Chip, a co-factor required for Apterous to function as a
transcription factor (Fernandez-Funez et
al., 1998
; Morcillo et al.,
1997
), or by expressing the Apterous antagonist dLMO
(Milán et al., 1998
).
In the D compartment, clones lacking Apterous activity were round in shape
with smooth borders (Fig. 4A
and see Fig. 5A). Clones mutant
for the apterous gene behaved similarly in the D compartment
(Blair et al., 1994
), but for
technical reasons were difficult to produce in large numbers (see Materials
and Methods for details).
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We measured the shape of wild-type clones and clones lacking Apterous
activity using the formula 4 A/L2 (A=area and L=perimeter of
the clone). 4
A/L2 equals 1.0 for a circle. The irregularly
shaped wild-type clones had a longer perimeter relative to their area and
generated a low value (4
A/L2=0.36;
Fig. 4A, quantitation in 4B). Dorsal clones lacking Apterous activity were significantly rounder than
wild-type clones (Chip mutant clones: 4
A/L2=0.68 and
P<<0.001; dLMO-expressing clones: 4
A/L2=0.81
and P<<0.001). Dorsal clones mutant for fringe were on
average rounder in shape than wild-type clones
(Fig. 4A; 4
A/L2=0.49 and P<0.001), but not as round as clones
lacking Apterous activity (P<0.001). This difference is reflected
in the observation that the borders of fringe mutant clones were
irregular and highly indented compared with the relatively smooth borders of
clones lacking Ap activity. These results agree with an earlier report that
loss of fringe activity has an effect on clone shape
(Rauskolb et al., 1999
).
However, our findings indicate that loss of fringe is not comparable
with loss of Apterous activity in the severity of its effect on the local cell
interactions that lead to sorting out of cells and smooth clone borders.
Comparable results were obtained when Apterous or Fringe was ectopically
expressed in the V compartment. Ventral Apterous-expressing clones were
significantly rounder than wild-type clones
(Fig. 4A and B; 4
A/L2=0.83 and P<<0.001). Ventral clones expressing
Fringe are on average rounder in shape than wild-type clones
(Fig. 4B; 4
A/L2=0.53 and P<0.001), but not as round as clones
expressing Apterous (P<0.001). These results indicate manipulation
of Apterous activity confers a larger difference in cell affinities than
modulation of Fringe alone can do, despite induction of Notch signaling.
Consistent with this conclusion, we find that expression of the constitutively
activated form of Notch in clones of cells is not sufficient to induce an
affinity difference in the wing pouch, reflected by failure of the clones to
adopt a round shape (Fig.
4C).
Apterous cannot confer a sustained affinity difference without Notch
activation
Dorsal clones lacking Apterous activity lose Fringe expression. Thus, it is
possible that the effects of clones lacking Apterous activity on cell affinity
may be due to loss of Fringe. The observation that removing Fringe activity
did not produce as robust an effect on clone shape as removing Apterous
activity suggested that Apterous would have multiple targets through which it
affects cell affinity, Fringe being one of these. In this case we would expect
that restoring Fringe activity in an apterous mutant clone would only
partially compensate for the cell affinity defects. To test this possibility,
we produced clones that lacked Apterous activity due to expression of the
inhibitor dLMO (Milán et al.,
1998). Use of Gal4 to remove Apterous activity allowed us to
evaluate the effects of restoring Fringe expression on the shape of clones
lacking expression of other Apterous target genes. Dorsal clones expressing
dLMO were round and had smooth borders
(Fig. 5A,E; 4
A/L2=0.81). When Fringe was co-expressed with dLMO, clones were
elongated in shape and had irregular borders
(Fig. 5B,E; 4
A/L2=0.34). As expected, Notch was not activated at the borders of
these clones and Wg expression was not induced. However, contrary to our
expectations, these clones were not significantly different in shape from
wild-type clones (P=0.7). This indicates that Apterous-dependent
alterations in cell affinity require the activity of Fringe, and therefore
presumably Notch activation.
To test the dependence of dLMO-expressing clone shape on Notch activity
more directly, we co-expressed a dominant-negative version of the Notch
receptor. Necd encodes a truncated form of Notch that lacks the
intracellular domain of the receptor, which blocks Notch activation in a
cell-autonomous manner (Micchelli and
Blair, 1999). When dLMO was used to remove Apterous activity,
Notch signaling was induced in cells on both sides of the border of the clone,
as revealed by Wg or Cut expression (Fig.
5A and data not shown, see drawing). This resembles the wild-type
DV border, in that feedback signaling leads to activation of Notch on both
sides of the interface between the two cell types
(de Celis and Bray, 1997
;
Micchelli et al., 1997
). Under
these conditions, Delta and Serrate would be induced in cells adjacent to the
Wg-expressing cells (i.e. offset from the borders of the clone by one or two
rows of cells). When Necd was co-expressed, we observed two
distinct outcomes. Notch activity and Wg and Cut expression were always lost
in the mutant cells (Fig. 5C,D;
see drawing). In some clones Notch was not activated in the adjacent wild-type
cells, and Wg and Cut failed to be induced. These clones were irregular in
shape (Fig. 5D,E; 4
A/L2=0.35). In other clones, Notch was activated and Wg and Cut
were expressed in surrounding cells. Clones of this type were round in shape
(Fig. 5C,E; 4
A/L2=0.77). When Notch activity was blocked in dLMO-expressing
clones by co-expression of Hairless or a dominant-negative form of Mastermind,
Wg and Cut failed to be induced inside the clone, but they were expressed in
surrounding cells (not shown). Clones of this type were round in shape, as
when Necd was co-expressed. The correlation between clone shape and
Notch signaling in adjacent cells allows the possibility that recruitment of
Serrate and Delta by Wg signaling may contribute to smoothing the clone
border. It is interesting to note that in clones with a round shape, only some
surrounding cells expressed Cut and Wg
(Fig. 5C). This observation
suggests that a lower level of Notch signaling activity is required to induce
the genes that confer the affinity difference than is required to induce
wg and cut expression.
Taken together, the data presented indicate that clones round up when Notch
is activated in cells outside the clone, but fail to do so when Notch is not
activated. This raises a question of why Necd is sometimes able to
block Notch activation in cells outside the clone. One possibility is that,
being a transmembrane protein, Necd may influence the ability of
Serrate, Delta or Notch to function in the neighboring cells (we cannot
exclude the possibility that Necd might also interact with other as
yet unidentified proteins to contribute to this effect). We note that
similarly variable effects were observed in Serrate-expressing clones
(Rauskolb et al., 1999). These
results suggest that the effects of Fringe can be fully accounted for in terms
of Notch activity. This contrasts with the previous proposal that Fringe acts
independently of Notch in DV cell affinity
(Rauskolb et al., 1999
;
O'Keefe and Thomas, 2001
).
Sorting out at the DV boundary
The DV compartment boundary behaves as a lineage restriction boundary.
Clones of cells born in one compartment do not give rise to progeny located in
the adjacent one (Fig. 6A,D).
This may be due to differences in affinity between D and V cells. However, the
DV compartment boundary behaves also as a signaling center. Interactions
between D and V cells induce Notch activation and Wg expression along the
boundary (Fig. 1A). As
illustrated in Fig. 6B, cells
that change affinity and also acquire the signaling properties of the opposite
compartment are expected to cross the boundary to intermingle freely with
cells in the new compartment. By contrast, acquisition of the signaling
properties of the opposite compartment might only be expected to cause
displacement of the Wg stripe without allowing the clone to cross completely
into the opposite compartment (Fig.
6C).
Apterous confers both the signaling and affinity properties of D cells. When Ap-expressing clones are produced in the V compartment, they cross into the D compartment and displace the Wg stripe (around the clone; Fig. 6E). The vast majority of these clones are topologically located in the D compartment, where they mix perfectly with D cells (Table 1). Likewise, clones lacking Ap activity in the D compartment because of dLMO expression can cross completely into the V compartment if they contact the DV boundary. All clones analyzed displace the Wg stripe and intermingle freely with V cells (Fig. 6F,G; Table 1).
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Fringe expression confers the signaling behavior of D cells, but as indicated above, appears not to confer all the affinity properties of D cells. We compared the behavior of Fringe-expressing clones born in the V compartment, with those of Ap. Although Fringe expressing clones displaced the Wg stripe with respect to the endogenous DV boundary in the same way as Ap-expressing clones (Fig. 6H), 6/10 clones crossed completely into the D compartment (Table 1). Co-expression of Serrate gave similar results; only 4/11 clones crossed completely into the D compartment. Likewise, fringe mutant clones born in the D compartment displaced the Wg stripe (Fig. 6I). However, only 9/23 clones were topologically located in the V compartment (Fig. 6I; Table 1). These findings provide another indication that Fringe is not comparable with Ap in establishing the cell affinities that control boundary formation.
As noted in the preceding section, Ap requires Notch activity to cause rounding up of clones. We therefore asked whether Notch activation is also required for sorting across the compartment boundary. We co-expressed Fringe to prevent activation of Notch in dLMO-expressing clones. In contrast to clones expressing dLMO alone, these clones did not sort into the V compartment (0/12 clones examined; Fig. 6J, compare with 6F,G). Comparable results were obtained when the dominant-negative form of the Notch receptor Necd was co-expressed with dLMO (Fig. 6K and K'). Clones that did not induce Notch activation in the adjacent wild-type cells did not sort out into the V compartment (Table 1).
Taken together, the results of the experiments with Fringe and Necd indicate that activation of Notch signaling is required for a sustained affinity difference between cells that express Apterous and those that do not. However, fringe cannot fully account for the effects of Apterous on D cell affinity. This comparison indicates that it is not the presence or absence of Fringe or of Notch activity per se that produces the affinity difference. Rather, Notch signaling appears to be required in conjunction with another Apterous-dependent process for maintenance of the affinity border.
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DISCUSSION |
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A second, very different, model proposes that Notch activation confers a
boundary-specific affinity state and that this is modulated into D and V
states by Apterous expression (Fig.
7C) (Micchelli and Blair,
1999; Blair, 2001
).
According to this model, there should be an affinity difference between
boundary cells and internal cells within a compartment but not between D and V
cells in the absence of Notch activity. This model proposes that Notch
activity is sufficient to produce an affinity difference and hence smooth
clone borders. However, as we have shown, clones of cells expressing the
activated Notch receptor do not exhibit this property
(Fig. 4C). This model is also
difficult to reconcile with our observation that the borders of
fringe mutant clones in the D compartment are highly irregular
(Fig. 4A; illustrated in
Fig. 7G). It is also
incompatible with our finding that restoring Notch activity in the absence of
Apterous function is not sufficient to generate a smooth DV boundary and
prevent mixing of D and V cells (Fig.
2; illustrated in
7E).
The results reported here support the view that Notch activity is needed
for cell affinity differences between D and V cells, but indicate that Notch
activation is not sufficient to cause these differences. We therefore propose
the model in Fig. 7D, which
differs in one crucial respect from the model discussed above
(Fig. 7C). We consider the role
of Notch activation to be permissive rather than instructive, and suggest that
Apterous controls expression of surface proteins in D and V cells. We envisage
that Notch activity is an essential co-factor in allowing cells to convert
this into an affinity state. In molecular terms, one possibility is that D and
V surface proteins form complexes with activated Notch (N*). In
this scenario D+N* and V+N* are the active components, D
and V are needed and instructive but have no activity alone. Interestingly, it
has been observed that loss of Notch activation only in one compartment does
not alter the DV affinity boundary
(Micchelli and Blair, 1999;
Milán and Cohen, 1999a
;
Rauskolb et al., 1999
). Thus,
production of either the dorsal (D+N*) or the ventral
(V+N*) boundary-specific cell state is sufficient to induce an
affinity difference with cells of the opposite compartment
(Fig. 7F). Another plausible
molecular scenario is that Notch activity might control the subcellular
localization of the predicted D and V proteins.
We present these examples to illustrate how Notch activity can be seen as a permissive co-factor rather than as an instructive principle defining cell affinity. Many other molecular explanations are possible. This model provides a satisfactory explanation for how Notch can be required, but not sufficient for boundary maintenance. The essential difference between the permissive and instructive models for Notch function lies in the observation that Notch activation leads to an affinity difference only in the context of juxtaposition of cells with opposite DV identity. Notch activation per se does not induce a robust affinity boundary, whereas clones expressing dLMO and Necd did so only when Notch was not blocked in the cells outside the clone (Fig. 7F). Comparable results were obtained with clones expressing Apterous and Necd.
Are the transmembrane proteins Serrate and Delta the D and V proteins,
respectively? Early in development, Serrate is expressed in D cells and Delta
in V cells (Diaz-Benjumea and Cohen,
1995; Milán and Cohen,
2000
) (Fig. 1A).
Late in development, both genes are regulated by Wg and are expressed in cells
adjacent to the Wg-expressing cells at the DV boundary
(de Celis and Bray, 1997
;
Micchelli et al., 1997
)
(Fig. 1B). Given that the
Serrate- and Delta-expressing cells are offset from the DV boundary, we
consider it unlikely that they confer the D* and V*
activities. However, we do not exclude the possibility that they might
contribute to the establishment of the DV affinity boundary in collaboration
with Caps and Tartan.
Cell behavior at the DV boundary: sorting out by crossing versus
pushing
The interface between D and V cells behaves as an affinity boundary and as
a signaling center where Notch activation is required for the growth of the
wing disc. Clones of cells can be induced to sort into the opposite
compartment by manipulating Apterous or Fringe activities. As discussed
previously by Blair and Ralston in the context of the AP boundary
(Blair and Ralston, 1997), we
would like to distinguish between crossing and pushing the DV boundary as
possible mechanisms. Cells with altered Apterous activity also have altered
Fringe activity. We suggest that these clones can cross the boundary and mix
freely with cells in the opposite compartment because they change both their
affinity state and signaling properties. Clones in which only Fringe activity
is altered adopt signaling properties of the opposite compartment and displace
the signaling center relative to the endogenous compartment boundary
(Fig. 7H)
(Rauskolb et al., 1999
). In
wild-type discs, symmetric activation of Notch and its targets leads to
symmetric growth of D and V compartments. If growth is symmetric with respect
to the displaced signaling center, the clone could be pushed into the opposite
compartment by growth of the surrounding tissue
(Blair and Ralston, 1997
) (see
also Fig. 7H).
At first glance, differential growth might explain how cells could be
pushed to the interface between compartments. Can the model presented in the
preceding section explain why some dorsal fringe mutant clones become
able to mix with cells of the opposite compartment? As shown by Rauskolb et
al. (Rauskolb et al., 1999)
Notch is not activated in V cells adjacent to fringe mutant clones
abutting the boundary (Fig.
7H). Our model suggests that these cells would become V instead of
V+N*; hence, there would not be a sustained affinity difference
between fringe mutant D cell and the adjacent V cells. This may
explain why fringe mutant D cells can sometimes mix with V cells when
they are pushed into the V compartment. A similar case can be made to explain
how V cells expressing Fringe can be pushed into the D compartment and mix
with D cells. In both situations, we note that these clones form smooth
borders with the cells of the compartment of origin, suggesting symmetric
growth induced by Notch may contribute to the smoothness of the affinity
boundary. This type of `pushing' mechanism provides a useful explanation for
the behavior of clones of cells that contact the DV boundary. We note that the
behavior of cells expressing Apterous and Fringe was not the same when the
entire P compartment was involved. P cells of ventral origin expressing
Apterous were able to sort into to dorsal posterior quadrant, but cells
expressing Fringe were not. We suggest that this reflects an underlying
difference between cells that have acquired a fully dorsal affinity state from
those in which only the signaling properties have been altered. Fringe
activity clearly plays an important role in the maintaining the segregation of
D and V cells, but it is not the sole mediator of Apterous activity in this
process.
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ACKNOWLEDGMENTS |
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REFERENCES |
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Blair, S. S. (1995). Compartments and appendage development in Drosophila. BioEssays 17,299 -309.[Medline]
Blair, S. S. (2001). Cell lineage: compartments and Capricious. Curr. Biol 11,R1017 -R1021.[CrossRef][Medline]
Blair, S. S., Brower, D. L., Thomas, J. B. and Zavortink, M.
(1994). The role of apterous in the control of
dorsoventral compartmentalization and PS integrin gene expression in the
developing wing of Drosophila. Development
120,1805
-1815.
Blair, S. S. and Ralston, A. (1997).
Smoothened-mediated Hedgehog signalling is required for the maintenance of the
anterior-posterior lineage restriction in the developing wing of Drosophila.
Development 124,4053
-4063.
Brückner, K., Perez, L., Clausen, H. and Cohen, S. M. (2000). Glycosytransferase activity of Fringe modulates Notch-Delta interactions. Nature 406,411 -415.[CrossRef][Medline]
Calleja, M., Moreno, E., Pelaz, S. and Morata, G.
(1996). Visualization of gene expression in living adult
Drosophila. Science 274,252
-255.
Cohen, B., McGuffin, M. E., Pfeifle, C., Segal, D. and Cohen, S. M. (1992). apterous: a gene required for imaginal disc development in Drosophila encodes a member of the LIM family of developmental regulatory proteins. Genes Dev. 6, 715-729.[Abstract]
de Celis, J. F. and Bray, S. (1997). Feed-back
mechanisms affecting Notch activation at the dorsoventral boundary in the
Drosophila wing. Development
124,3241
-3251.
de Celis, J. F., Garcia-Bellido, A. and Bray, S. J.
(1996). Activation and function of Notch at the
dorsal-ventral boundary of the wing imaginal disc.
Development 122,359
-369.
Diaz-Benjumea, F. J. and Cohen, S. M. (1995).
Serrate signals through Notch to establish a Wingless-dependent organizer at
the dorsal/ventral compartment boundary of the Drosophila wing.
Development 121,4215
-4225.
Doherty, D., Fenger, G., Younger-Shepherd, S., Jan, L.-Y. and Jan, Y.-N. (1996). Dorsal and ventral cells respond differently to the Notch ligands Delta and Serrate during Drosophila wing development. Genes Dev. 10,421 -434.[Abstract]
Fernández-Funez, P., Lu, C. H., Rincon-Limas, D. E.,
Garcia-Bellido, A. and Botas, J. (1998). The relative
expression amounts of apterous and its co-factor dLdb/Chip are critical for
dorso-ventral compartmentalization in the Drosophila wing. EMBO
J. 17,6846
-6853.
Fietz, M. J., Jacinto, A., Taylor, A. M., Alexandre, C. and Ingham, P. W. (1995). Secretion of the amino-terminal fragment of Hedgehog protein is necessary and sufficient for hedgehog signalling in Drosophila. Curr. Biol. 5, 643-650.[Medline]
Fleming, R. J., Gu, Y. and Hukriede, N. A.
(1997). Serrate-mediated activation of Notch is specifically
blocked by the product of the gene fringe in the dorsal compartment of the
Drosophila wing imaginal disc. Development
124,2973
-2981.
García-Bellido, A., Ripoll, P. and Morata, G. (1973). Developmental compartmentalisation of the wing disk of Drosophila. Nature 245,251 -253.
Giraldez, A. J., Perez, L. and Cohen, S. M. (2002). A naturally occurring alternative product of the mastermind locus that represses notch signalling. Mech. Dev. 115,101 -105.[CrossRef][Medline]
Go, M. J., Eastman, D. S. and Artavanis-Tsakonas, S.
(1998). Cell proliferation control by Notch signaling in
Drosophila development. Development
125,2031
-2040.
Irvine, K. and Wieschaus, E. (1994). fringe, a boundary specific signalling molecule, mediates interactions between dorsal and ventral cells during Drosophila wing development. Cell 79,595 -606.[Medline]
Lawrence, P. A., Casal, J. and Struhl, G.
(1999). The Hedgehog morphogen and gradients of cell affinity in
the abdomen of Drosophila. Development
126,2441
-2449.
Lecuit, T., Brook, W. J., Ng, M., Calleja, M., Sun, H. and Cohen, S. M. (1996). Two distinct mechanisms for long-range patterning by Decapentaplegic in the Drosophila wing. Nature 381,387 -393.[CrossRef][Medline]
Liu, X., Grammont, M. and Irvine, K. D. (2000). Roles for scalloped and vestigial in regulating cell affinity and interactions between the wing blade and the wing hinge. Dev. Biol. 228,287 -303.[CrossRef][Medline]
Micchelli, C. A. and Blair, S. S. (1999). Dorsoventral lineage restriction in wing imaginal discs requires Notch. Nature 401,473 -476.[CrossRef][Medline]
Micchelli, C. A., Rulifson, E. J. and Blair, S. S.
(1997). The function and regulation of cut expression on the wing
margin of Drosophila: Notch, Wingless and a dominant negative role for Delta
and Serrate. Development
124,1485
-1495.
Milán, M. and Cohen, S. M. (1999a). Notch signaling is not sufficient to define the affinity boundary between dorsal and ventral compartments. Mol. Cell 4,1073 -1078.[Medline]
Milán, M. and Cohen, S. M. (1999b). Regulation of LIM homeodomain activity in vivo: A tetramer of dLDB and Apterous confers activity and capacity for regulation by ddLMO. Mol. Cell 4,267 -273.[Medline]
Milán, M. and Cohen, S. M. (2000).
Temporal regulation of Apterous activity during development of the Drosophila
wing. Development 127,3069
-3078.
Milán, M., Diaz-Benjumea, F. and Cohen, S. M.
(1998). Beadex encodes an dLMO protein that regulates
Apterous LIM-homeodomain activity in Drosophila wing development: a
model for dLMO oncogene function. Genes Dev.
12,2912
-2920.
Milán, M., Weihe, U., Perez, L. and Cohen, S. M. (2001). The LRR proteins capricious and Tartan mediate cell interactions during DV boundary formation in the Drosophila wing. Cell 106,785 -794.[Medline]
Milán, M., Pérez, L. and Cohen, S. M. (2002). Short-range cell interactions and cell survival in the Drosophila wing. Dev. Cell 2, 797-805.[Medline]
Moloney, D. J., Panin, V. M., Johnston, S. H., Chen, J., Shao, L., Wilson, R., Wang, Y., Stanley, P., Irvine, K. D., Haltiwanger, R. S. et al. (2000). Fringe is a glycosyltransferase that modifies Notch. Nature 406,369 -375.[CrossRef][Medline]
Morcillo, P., Rosen, C., Baylies, M. K. and Dorsett, D.
(1997). Chip, a widely expressed chromosomal protein required for
segmentation and activity of a remote wing margin enhancer in Drosophila.Genes Dev. 11,2729
-2740.
Munro, S. and Freeman, M. (2000). The notch signalling regulator fringe acts in the Golgi apparatus and requires the glycosyltransferase signature motif DXD. Curr. Biol. 10,813 -820.[CrossRef][Medline]
Nellen, D., Burke, R., Struhl, G. and Basler, K. (1996). Direct and long-range action of a DPP morphogen gradient. Cell 85,357 -368.[Medline]
Neumann, C. J. and Cohen, S. M. (1997).
Long-range action of Wingless organizes the dorsal-ventral axis of the
Drosophila wing. Development
124,871
-880.
O'Keefe, D. D. and Thomas, J. B. (2001).
Drosophila wing development in the absence of dorsal identity.
Development 128,703
-710.
Panin, V. M., Papayannopoulos, V., Wilson, R. and Irvine, K. D. (1997). Fringe modulates Notch-ligand interactions. Nature 387,908 -913.[CrossRef][Medline]
Pignoni, F. and Zipursky, S. L. (1997).
Induction of Drosophila eye development by decapentaplegic.
Development 124,271
-278.
Rauskolb, C., Correia, T. and Irvine, K. D. (1999). Fringe-dependent separation of dorsal and ventral cells in the Drosophila wing. Nature 401,476 -480.[CrossRef][Medline]
Rodriguez, I. and Basler, K. (1997). Control of compartmental affinity boundaries by hedgehog. Nature 389,614 -618.[CrossRef][Medline]
Weihe, U., Milán, M. and Cohen, S. M.
(2001). Regulation of Apterous activity in Drosophila wing
development. Development
128,4615
-4622.
Zecca, M., Basler, K. and Struhl, G. (1996). Direct and long-range action of a Wingless morphogen gradient. Cell 87,833 -844.[Medline]