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First published online 19 July 2006
doi: 10.1242/dev.02492

Development 133, 3255-3264 (2006)
Published by The Company of Biologists 2006
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Differential expression of the adhesion molecule Echinoid drives epithelial morphogenesis in Drosophila

Caroline Laplante and Laura A. Nilson*

Department of Biology, McGill University, 1205 Doctor Penfield Avenue, Montréal, QC H3A 1B1, Canada.


Figure 1
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  		Fig. 1. F72 homozygous follicle cell clones produce groups of eggshell imprints with a smooth border. In all figures, anterior is towards the left and dorsal is towards the top, unless otherwise indicated. (A) Wild-type egg. (B) Egg from an F72 mosaic female. Groups of surface imprints exhibit smooth borders (arrows). (C) Egg from a female with homozygous F72 clones marked with the dec-1 eggshell marker (arrows). (D) Egg from an edlF20 mosaic female.(E,E') Clone of F72 homozygous follicle cells in a stage 11 egg chamber. (E) Clone is marked by the absence of the NM clone marker (basal confocal section); heterozygous and homozygous wild type cells are also visible. (E') Anti-phosphotyrosine (p-Tyr) staining, apical confocal section. The apical clone border is smooth. (F,F') Rhodamine-phalloidin staining to visualize filamentous actin (F-actin). F72 mutant clones at stage 10B (F, basal confocal section) do not exhibit a smooth border (F', apical confocal section).

 

Figure 2
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  		Fig. 2. ed clone borders induce the formation of a contractile actin cable and reduction in adherens junction components. (A) edF72 follicle cell clone (lack of NM marker, basal confocal section). (A') Staining with fluorescently-labeled phalloidin (apical confocal section) reveals enriched F-actin at the clone border and reduced apical circumference.(B,B') Cross-section of an edF72 mutant clone (B) stained with fluorescently-labeled phalloidin (B') illustrates the apical constriction of the clone border (arrows, apical is towards the bottom). (C) edF72 follicle cell clone (basal confocal section). (C') Increased p-MLC immunoreactivity (apical confocal section) is detected at the clone border.(D,D') Enlargement of an edF72 clone border. Two parallel lines of p-MLC immunoreactivity can be resolved. (E,E') Stage 12. Some edF72 mutantclones exhibit severely reduced or discontinuous DE-cad immunoreactivity at the clone border (arrow). (F,F') Stage 12. At edF72 mutant clone borders with a milder DE-Cad defect, DE-Cad is occasionally absent (arrow) but often discontinuous or unaffected (arrowhead).

 

Figure 3
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  		Fig. 3. Ed exhibits a spatially and temporally dynamic distribution during oogenesis. (A) Homozygous edF72 follicle cell clone (lack of NM marker, basal confocal section). (A') Ed immunoreactivity is absent from the ed mutant cells and absent (arrow) or discontinuous (arrowhead) from wild-type cells at the clone border (apical confocal section). (B) Ed is enriched at the apical (germline-facing) side of the tissue (inset; apical is indicated with an arrowhead, basal with an arrow) at early stages (left) then becomes undetectable by early stage 10B (right). (C) Late stage 10B (dorsal view). Ed is detected at the dorsal midline of the main body follicle cells (only the posterior half of the egg chamber is shown). (C') Same egg chamber as in C, labeled with fluorescently-labeled phalloidin. A smooth interface coincides with the endogenous Ed expression border (arrow). (D) Stage 11 (dorsolateral view). Ed is absent from two dorsolateral groups of follicle cells. (D') Same egg chamber as in D (dorsal anterior portion), labeled with fluorescently-labeled phalloidin. A smooth interface (arrow, arrowhead) corresponds to the endogenous Ed expression border.

 

Figure 4
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  		Fig. 4. Absence of Ed from the roof cells generates an endogenous Ed expression border. (A,B) Cross-sections (lateral view) through the dorsal anterior follicular epithelium at early (A) and late (B) stage 11. Ed (red) is detected in cells expressing the rho-lacZ floor cell marker (green). The floor cell shown in A has begun to elongate posteriorly; this elongation is more pronounced by the stage shown in B. Nuclei are shown in blue. (C-C'') Stage 11 (dorsal view).(C) High Broad levels mark the roof cell nuclei. (C') Ed is present in all follicle cells except for two dorsolateral domains. (C'') Merge. Ed (red) is absent from the roof cells (green); one roof cell domain is outlined. (D-D'') Stage 11 (dorsal view). The two L-shaped floor cell domains (D, basal confocal section) align with the limit of the Ed domain (D', apical confocal section). (D'') Merge of Ed (red) and rho-lacZ (green). The patterns in merged images (C'',D'') are slightly out of register owing to the different planes of the images and the onset of morphogenesis (see A).

 

Figure 5
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  		Fig. 5. Ed is required for tube floor closure during appendage morphogenesis. (A) Stage 11 mosaic egg chamber (dorsal view) with a large ed mutant clone (lack of NM marker) that includes much of both appendage primordia. (A') Ed expression is detectable in the posterior non-mutant follicle cells (arrow), indicating that this egg chamber is of a stage at which Ed would be expressed in all follicle cells except the presumptive roof cells (see Fig. 3D). (A'') Anti-phosphotyrosine (p-Tyr) staining. The interface between the roof and floor cell domains is morphologically distinguishable but not smooth (arrow), and the posterior border of the roof cell domain (arrowhead) is not distinguishable. (B) Stage 12 egg chamber (dorsal view) with one wild-type (bottom) and one mutant (top, lack of NM marker) appendage primordium. (B',B'') Staining with fluorescently-labeled phalloidin, basal section (B'), apical confocal section (B''). The opening of the mutant tube appears wider (arrows). (C-C'') Diagram of a single appendage primordium at successive stages of tube formation. Both cross-section (left) and surface views (right; anterior towards the left, dorsal towards the top) are shown. The presumptive roof cells (light gray) are flanked anteriorly and medially by a single row of floor cells (dark gray). To emphasize the floor cell movements, roof cells are not delineated individually in the surface views. (D-I) Appendage primordia expressing the rho-lacZ floor cell marker. (D-F) Wild-type appendage primordia. (D) Primordium at onset of tube extension phase (floor cell domain aligned with oocyte/nurse cell margin). The floor cells have elongated and the apices nearest the `hinge' between the anterior and medial domains have met. (E) Tube extension phase (floor cell domain overlaps nurse cells). The tube floor is nearly fully closed. (F) A later stage than E. The tube floor is closed. (G-I) ed mutant primordia. (G) Same stage as D. The floor cell apices have not met. (H) Tube extension stage, comparable with E. The floor cell apices have not met; the tube floor remains open. (I) Later stage than H. The tube floor remains open. (J,J') Two sides of a single stage 14 egg chamber in which all follicle cells are mutant for ed. Autofluorescence and DAPI staining reveal the eggshell structure and nuclei of the follicular epithelium, respectively. The dorsal appendages are severely reduced (arrow).

 

Figure 6
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  		Fig. 6. Ed is required for dorsal closure. (A,A') Stage 14 embryo, dorsal view, zippering phase of dorsal closure (Jacinto et al., 2002Go). (A) Arm immunoreactivity highlights the amnioserosa (arrow) and surrounding lateral epidermis. (A') Ed immunoreactivity is absent from the amnioserosa (arrow). (B,C) Arm staining of edMZ mutant embryos at the zippering phase (B) and termination phase (C) of dorsal closure. There are gaps along the dorsal midline (arrow) and misaligned segments (arrowheads). (D-D'') Successive images of a live wild-type embryo expressing GFP-moesin during zippering (D,D') and termination (D''). Accumulation of GFP-moesin at the leading edge highlights the contractile cable (D,D', arrows). (E-E'') Live imaging of dorsal closure in an edMZ embryo expressing GFP-moesin. GFP-moesin does not accumulate at the leading edge (E,E', arrows), and the gaps and segments are misaligned in the termination phase (E'').

 

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© The Company of Biologists Ltd 2006