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Adherens junction-dependent and -independent steps in the establishment of epithelial cell polarity in Drosophila.

Harris TJ, Peifer M - J. Cell Biol. (2004)

Bottom Line: We found apical accumulation of both Drosophila E-Cadherin (DE-Cad) and the apical cue Bazooka (Baz) as cells first form.Some epithelial structures are retained, however.These structures maintain apical Baz, accumulate apical Crumbs, and organize polarized cytoskeletons, but display abnormal cell morphology and fail to segregate the basolateral cue Discs large from the apical domain.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.

ABSTRACT
Adherens junctions (AJs) are thought to be key landmarks for establishing epithelial cell polarity, but the origin of epithelial polarity in Drosophila remains unclear. Thus, we examined epithelial polarity establishment during early Drosophila development. We found apical accumulation of both Drosophila E-Cadherin (DE-Cad) and the apical cue Bazooka (Baz) as cells first form. Mutant analyses revealed that apical Baz accumulations can be established in the absence of AJs, whereas assembly of apical DE-Cad complexes requires Baz. Thus, Baz acts upstream of AJs during epithelial polarity establishment. During gastrulation the absence of AJs results in widespread cell dissociation and depolarization. Some epithelial structures are retained, however. These structures maintain apical Baz, accumulate apical Crumbs, and organize polarized cytoskeletons, but display abnormal cell morphology and fail to segregate the basolateral cue Discs large from the apical domain. Thus, although epithelial polarity develops in the absence of AJs, AJs play specific roles in maintaining epithelial architecture and segregating basolateral cues.

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armm/z mutants retain residual epithelial structure and cell polarity. (A–H′) WT gastrulation. (A) Honeycomb Baz pattern in stage 7 ectoderm. (B) Apical Baz along stage 7 epithelial furrows and invaginations (arrow; cephalic furrow). (C) Baz at ectodermal spot junctions at early gastrulation (arrow), and (D) belt junctions by late gastrulation. (E) Cross section. Dlg (green) overlaps Baz (red) and DE-Cad (blue) in spot junctions (arrow) at early gastrulation. (F) Cross section. Dlg (green) segregates from belt junctions by late gastrulation (arrow). (G–H′) At stage 7 Dlg (green) segregates from Baz (red; inset, arrow) and the apical domain in cephalic furrows (G and G′, arrows; inset is a close up) and PMGI (H and H′, arrows). (I–O) armm/z. (I) Honeycomb Baz pattern is retained in stage 7-8 ectoderm (staged by morphology). (J) Apical Baz in stage 7-8 epithelial folds (fold outlined). (K) Honeycomb Baz pattern is lost in stage 9-10 ectoderm (staged by zygotically rescued siblings). Note Baz on inward membranes of rosettes (outlined). (L) Apical Baz in stage 9-10 fragmented folds (outlined). (M and M′, inset is a close-up) Stage 7-8 armm/z early fold. Dlg (green) overlaps with Baz (red; arrows) and apical domain (blue arrowheads). Fold and single cell outlined; b, basal. (N and N′) Rosettes. Dlg (green) is in apical domain (blue arrowheads), but shows some segregation from Baz (red, arrow). Rosette outlined; b, basal. (O) Stage 9-10 armm/z mutant. In dissociated cells Dlg segregates from membrane domains exposed at the embryo surface (white arrowheads), but is around the entire cortex of internal dissociated cells (blue arrowhead). (P) Schematic of Baz at apex in WT epithelial cell and at collar in armm/z epithelial cells. (Q, Q′, and Q′′) Stage 7-8 shgm/z early fold. Dlg (green) shows some segregation from Baz (red; arrows) but mislocalizes to the apical domain (blue arrowhead). DE-Cad (gray) shows some accumulation in proximity to Baz. Single cell outlined; b, basal. (R–U) Constant confocal settings. Lower DE-Cad levels in stage 7-8 and 9-10 armm/z mutants (S and U) versus stage 7 and 9 zygotically rescued (maternally mutant) siblings (armm; R and T). (V) DE-Cad (green) and Baz (red) in early armm/z fold. Note minimal colocalization (arrow). (W) DE-Cad (green), (X) Arm (green), and (Y) α-cat (green) with Baz (red) in later armm/z rosettes. DE-Cad (W) and Arm (X) localize near Baz accumulations (arrows), but α-cat does not (Y; arrows). In images of AJ proteins in armm/z and shgm/z mutants, darker grays were converted to black to detect any AJ protein accumulation by removing the low level cytoplasmic staining. Bars: (gray) 25 μm; (white) 5 μm.
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fig6: armm/z mutants retain residual epithelial structure and cell polarity. (A–H′) WT gastrulation. (A) Honeycomb Baz pattern in stage 7 ectoderm. (B) Apical Baz along stage 7 epithelial furrows and invaginations (arrow; cephalic furrow). (C) Baz at ectodermal spot junctions at early gastrulation (arrow), and (D) belt junctions by late gastrulation. (E) Cross section. Dlg (green) overlaps Baz (red) and DE-Cad (blue) in spot junctions (arrow) at early gastrulation. (F) Cross section. Dlg (green) segregates from belt junctions by late gastrulation (arrow). (G–H′) At stage 7 Dlg (green) segregates from Baz (red; inset, arrow) and the apical domain in cephalic furrows (G and G′, arrows; inset is a close up) and PMGI (H and H′, arrows). (I–O) armm/z. (I) Honeycomb Baz pattern is retained in stage 7-8 ectoderm (staged by morphology). (J) Apical Baz in stage 7-8 epithelial folds (fold outlined). (K) Honeycomb Baz pattern is lost in stage 9-10 ectoderm (staged by zygotically rescued siblings). Note Baz on inward membranes of rosettes (outlined). (L) Apical Baz in stage 9-10 fragmented folds (outlined). (M and M′, inset is a close-up) Stage 7-8 armm/z early fold. Dlg (green) overlaps with Baz (red; arrows) and apical domain (blue arrowheads). Fold and single cell outlined; b, basal. (N and N′) Rosettes. Dlg (green) is in apical domain (blue arrowheads), but shows some segregation from Baz (red, arrow). Rosette outlined; b, basal. (O) Stage 9-10 armm/z mutant. In dissociated cells Dlg segregates from membrane domains exposed at the embryo surface (white arrowheads), but is around the entire cortex of internal dissociated cells (blue arrowhead). (P) Schematic of Baz at apex in WT epithelial cell and at collar in armm/z epithelial cells. (Q, Q′, and Q′′) Stage 7-8 shgm/z early fold. Dlg (green) shows some segregation from Baz (red; arrows) but mislocalizes to the apical domain (blue arrowhead). DE-Cad (gray) shows some accumulation in proximity to Baz. Single cell outlined; b, basal. (R–U) Constant confocal settings. Lower DE-Cad levels in stage 7-8 and 9-10 armm/z mutants (S and U) versus stage 7 and 9 zygotically rescued (maternally mutant) siblings (armm; R and T). (V) DE-Cad (green) and Baz (red) in early armm/z fold. Note minimal colocalization (arrow). (W) DE-Cad (green), (X) Arm (green), and (Y) α-cat (green) with Baz (red) in later armm/z rosettes. DE-Cad (W) and Arm (X) localize near Baz accumulations (arrows), but α-cat does not (Y; arrows). In images of AJ proteins in armm/z and shgm/z mutants, darker grays were converted to black to detect any AJ protein accumulation by removing the low level cytoplasmic staining. Bars: (gray) 25 μm; (white) 5 μm.

Mentions: Next, we tested whether the apical Baz accumulations established during armm/z mutant cellularization are maintained during gastrulation, and whether further polarity is established, specifically addressing the segregation of Dlg from the apical domain. During WT gastrulation and subsequent development, Baz encircles the apical domain of ectodermal cells in a honeycomb pattern (Fig. 6 A), and accumulates in the apical domain of epithelial folds (Fig. 6 B, arrow). During early gastrulation, Baz associates with apical spot junctions (Fig. 6 C, arrow), which coalesce into belt junctions by late gastrulation (Fig. 6 D). As noted above, the basolateral cue Dlg overlaps with apical spot junctions at the end of cellularization (Fig. 1 D′). This overlap continues during early gastrulation (Fig. 6 E, arrow), but Dlg segregates from the apical complexes after they fuse into belt junctions (Fig. 6 F, arrow). Baz and Dlg also segregate in early epithelial folds. In the early cephalic furrow, Baz localizes apically (Fig. 6 G, arrow) and Dlg segregates to the basolateral domain (Fig. 6 G′), showing enrichment just below apical Baz (Fig. 6 G, inset, arrow; some Dlg is seen in the apical cytoplasm). In the early PMGI, Baz also localizes to the cell apex, despite its constricted structure, and Dlg segregates to the basolateral domain (Fig. 6, H and H′, arrows).


Adherens junction-dependent and -independent steps in the establishment of epithelial cell polarity in Drosophila.

Harris TJ, Peifer M - J. Cell Biol. (2004)

armm/z mutants retain residual epithelial structure and cell polarity. (A–H′) WT gastrulation. (A) Honeycomb Baz pattern in stage 7 ectoderm. (B) Apical Baz along stage 7 epithelial furrows and invaginations (arrow; cephalic furrow). (C) Baz at ectodermal spot junctions at early gastrulation (arrow), and (D) belt junctions by late gastrulation. (E) Cross section. Dlg (green) overlaps Baz (red) and DE-Cad (blue) in spot junctions (arrow) at early gastrulation. (F) Cross section. Dlg (green) segregates from belt junctions by late gastrulation (arrow). (G–H′) At stage 7 Dlg (green) segregates from Baz (red; inset, arrow) and the apical domain in cephalic furrows (G and G′, arrows; inset is a close up) and PMGI (H and H′, arrows). (I–O) armm/z. (I) Honeycomb Baz pattern is retained in stage 7-8 ectoderm (staged by morphology). (J) Apical Baz in stage 7-8 epithelial folds (fold outlined). (K) Honeycomb Baz pattern is lost in stage 9-10 ectoderm (staged by zygotically rescued siblings). Note Baz on inward membranes of rosettes (outlined). (L) Apical Baz in stage 9-10 fragmented folds (outlined). (M and M′, inset is a close-up) Stage 7-8 armm/z early fold. Dlg (green) overlaps with Baz (red; arrows) and apical domain (blue arrowheads). Fold and single cell outlined; b, basal. (N and N′) Rosettes. Dlg (green) is in apical domain (blue arrowheads), but shows some segregation from Baz (red, arrow). Rosette outlined; b, basal. (O) Stage 9-10 armm/z mutant. In dissociated cells Dlg segregates from membrane domains exposed at the embryo surface (white arrowheads), but is around the entire cortex of internal dissociated cells (blue arrowhead). (P) Schematic of Baz at apex in WT epithelial cell and at collar in armm/z epithelial cells. (Q, Q′, and Q′′) Stage 7-8 shgm/z early fold. Dlg (green) shows some segregation from Baz (red; arrows) but mislocalizes to the apical domain (blue arrowhead). DE-Cad (gray) shows some accumulation in proximity to Baz. Single cell outlined; b, basal. (R–U) Constant confocal settings. Lower DE-Cad levels in stage 7-8 and 9-10 armm/z mutants (S and U) versus stage 7 and 9 zygotically rescued (maternally mutant) siblings (armm; R and T). (V) DE-Cad (green) and Baz (red) in early armm/z fold. Note minimal colocalization (arrow). (W) DE-Cad (green), (X) Arm (green), and (Y) α-cat (green) with Baz (red) in later armm/z rosettes. DE-Cad (W) and Arm (X) localize near Baz accumulations (arrows), but α-cat does not (Y; arrows). In images of AJ proteins in armm/z and shgm/z mutants, darker grays were converted to black to detect any AJ protein accumulation by removing the low level cytoplasmic staining. Bars: (gray) 25 μm; (white) 5 μm.
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Related In: Results  -  Collection

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fig6: armm/z mutants retain residual epithelial structure and cell polarity. (A–H′) WT gastrulation. (A) Honeycomb Baz pattern in stage 7 ectoderm. (B) Apical Baz along stage 7 epithelial furrows and invaginations (arrow; cephalic furrow). (C) Baz at ectodermal spot junctions at early gastrulation (arrow), and (D) belt junctions by late gastrulation. (E) Cross section. Dlg (green) overlaps Baz (red) and DE-Cad (blue) in spot junctions (arrow) at early gastrulation. (F) Cross section. Dlg (green) segregates from belt junctions by late gastrulation (arrow). (G–H′) At stage 7 Dlg (green) segregates from Baz (red; inset, arrow) and the apical domain in cephalic furrows (G and G′, arrows; inset is a close up) and PMGI (H and H′, arrows). (I–O) armm/z. (I) Honeycomb Baz pattern is retained in stage 7-8 ectoderm (staged by morphology). (J) Apical Baz in stage 7-8 epithelial folds (fold outlined). (K) Honeycomb Baz pattern is lost in stage 9-10 ectoderm (staged by zygotically rescued siblings). Note Baz on inward membranes of rosettes (outlined). (L) Apical Baz in stage 9-10 fragmented folds (outlined). (M and M′, inset is a close-up) Stage 7-8 armm/z early fold. Dlg (green) overlaps with Baz (red; arrows) and apical domain (blue arrowheads). Fold and single cell outlined; b, basal. (N and N′) Rosettes. Dlg (green) is in apical domain (blue arrowheads), but shows some segregation from Baz (red, arrow). Rosette outlined; b, basal. (O) Stage 9-10 armm/z mutant. In dissociated cells Dlg segregates from membrane domains exposed at the embryo surface (white arrowheads), but is around the entire cortex of internal dissociated cells (blue arrowhead). (P) Schematic of Baz at apex in WT epithelial cell and at collar in armm/z epithelial cells. (Q, Q′, and Q′′) Stage 7-8 shgm/z early fold. Dlg (green) shows some segregation from Baz (red; arrows) but mislocalizes to the apical domain (blue arrowhead). DE-Cad (gray) shows some accumulation in proximity to Baz. Single cell outlined; b, basal. (R–U) Constant confocal settings. Lower DE-Cad levels in stage 7-8 and 9-10 armm/z mutants (S and U) versus stage 7 and 9 zygotically rescued (maternally mutant) siblings (armm; R and T). (V) DE-Cad (green) and Baz (red) in early armm/z fold. Note minimal colocalization (arrow). (W) DE-Cad (green), (X) Arm (green), and (Y) α-cat (green) with Baz (red) in later armm/z rosettes. DE-Cad (W) and Arm (X) localize near Baz accumulations (arrows), but α-cat does not (Y; arrows). In images of AJ proteins in armm/z and shgm/z mutants, darker grays were converted to black to detect any AJ protein accumulation by removing the low level cytoplasmic staining. Bars: (gray) 25 μm; (white) 5 μm.
Mentions: Next, we tested whether the apical Baz accumulations established during armm/z mutant cellularization are maintained during gastrulation, and whether further polarity is established, specifically addressing the segregation of Dlg from the apical domain. During WT gastrulation and subsequent development, Baz encircles the apical domain of ectodermal cells in a honeycomb pattern (Fig. 6 A), and accumulates in the apical domain of epithelial folds (Fig. 6 B, arrow). During early gastrulation, Baz associates with apical spot junctions (Fig. 6 C, arrow), which coalesce into belt junctions by late gastrulation (Fig. 6 D). As noted above, the basolateral cue Dlg overlaps with apical spot junctions at the end of cellularization (Fig. 1 D′). This overlap continues during early gastrulation (Fig. 6 E, arrow), but Dlg segregates from the apical complexes after they fuse into belt junctions (Fig. 6 F, arrow). Baz and Dlg also segregate in early epithelial folds. In the early cephalic furrow, Baz localizes apically (Fig. 6 G, arrow) and Dlg segregates to the basolateral domain (Fig. 6 G′), showing enrichment just below apical Baz (Fig. 6 G, inset, arrow; some Dlg is seen in the apical cytoplasm). In the early PMGI, Baz also localizes to the cell apex, despite its constricted structure, and Dlg segregates to the basolateral domain (Fig. 6, H and H′, arrows).

Bottom Line: We found apical accumulation of both Drosophila E-Cadherin (DE-Cad) and the apical cue Bazooka (Baz) as cells first form.Some epithelial structures are retained, however.These structures maintain apical Baz, accumulate apical Crumbs, and organize polarized cytoskeletons, but display abnormal cell morphology and fail to segregate the basolateral cue Discs large from the apical domain.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.

ABSTRACT
Adherens junctions (AJs) are thought to be key landmarks for establishing epithelial cell polarity, but the origin of epithelial polarity in Drosophila remains unclear. Thus, we examined epithelial polarity establishment during early Drosophila development. We found apical accumulation of both Drosophila E-Cadherin (DE-Cad) and the apical cue Bazooka (Baz) as cells first form. Mutant analyses revealed that apical Baz accumulations can be established in the absence of AJs, whereas assembly of apical DE-Cad complexes requires Baz. Thus, Baz acts upstream of AJs during epithelial polarity establishment. During gastrulation the absence of AJs results in widespread cell dissociation and depolarization. Some epithelial structures are retained, however. These structures maintain apical Baz, accumulate apical Crumbs, and organize polarized cytoskeletons, but display abnormal cell morphology and fail to segregate the basolateral cue Discs large from the apical domain. Thus, although epithelial polarity develops in the absence of AJs, AJs play specific roles in maintaining epithelial architecture and segregating basolateral cues.

Show MeSH
Related in: MedlinePlus