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The positioning and segregation of apical cues during epithelial polarity establishment in Drosophila.

Harris TJ, Peifer M - J. Cell Biol. (2005)

Bottom Line: Adherens junctions (AJs) often direct this polarity, but we previously found that Bazooka (Baz) acts upstream of AJs as epithelial polarity is first established in Drosophila.Surprisingly, we found that Baz localizes to an apical domain below its typical binding partners atypical protein kinase C (aPKC) and partitioning defective (PAR)-6 as the Drosophila epithelium first forms.These results reveal key steps in the assembly of the apical domain in Drosophila.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA. tonyh@email.unc.edu

ABSTRACT
Cell polarity is critical for epithelial structure and function. Adherens junctions (AJs) often direct this polarity, but we previously found that Bazooka (Baz) acts upstream of AJs as epithelial polarity is first established in Drosophila. This prompted us to ask how Baz is positioned and how downstream polarity is elaborated. Surprisingly, we found that Baz localizes to an apical domain below its typical binding partners atypical protein kinase C (aPKC) and partitioning defective (PAR)-6 as the Drosophila epithelium first forms. In fact, Baz positioning is independent of aPKC and PAR-6 relying instead on cytoskeletal cues, including an apical scaffold and dynein-mediated basal-to-apical transport. AJ assembly is closely coupled to Baz positioning, whereas aPKC and PAR-6 are positioned separately. This forms a stratified apical domain with Baz and AJs localizing basal to aPKC and PAR-6, and we identify specific mechanisms that keep these proteins apart. These results reveal key steps in the assembly of the apical domain in Drosophila.

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A specific role for dynein in positioning Baz and AJs. dhc64Cm/z mutants. (A–C) Cellularization. (A) Baz (red) and DE-Cad (green) mislocalize basally but often colocalize. WT, right. (B) aPKC (red) enriched apically. DE-Cad mislocalized basally (green). (C) PAR-6 (green) localizes as in WT. (D–G) Gastrulation. (D) Baz (red) and DE-Cad (green) are apical and colocalize. (E) aPKC apical. (F) PAR-6 apical. (G) Crb apical. (H and I) PMGIs. (H) aPKC (red) apical to DE-Cad (green) and Dlg (blue) segregates basally. (I) PAR-6 (green) above Baz (red) and Dlg (blue) basal. Bars, 5 μm.
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fig6: A specific role for dynein in positioning Baz and AJs. dhc64Cm/z mutants. (A–C) Cellularization. (A) Baz (red) and DE-Cad (green) mislocalize basally but often colocalize. WT, right. (B) aPKC (red) enriched apically. DE-Cad mislocalized basally (green). (C) PAR-6 (green) localizes as in WT. (D–G) Gastrulation. (D) Baz (red) and DE-Cad (green) are apical and colocalize. (E) aPKC apical. (F) PAR-6 apical. (G) Crb apical. (H and I) PMGIs. (H) aPKC (red) apical to DE-Cad (green) and Dlg (blue) segregates basally. (I) PAR-6 (green) above Baz (red) and Dlg (blue) basal. Bars, 5 μm.

Mentions: To directly test whether dynein functions in positioning Baz, we analyzed the Baz distribution in cellularizing dhc64Cm/z mutants. These mutants have defects in syncytial nuclear divisions (Robinson et al., 1999), but many undergo partial cellularization and initiate morphogenesis. In the mutants, centrosomes are positioned apically during cellularization (unpublished data), suggesting that basic MT polarity is retained. Baz and DE-Cad are recruited to early cellularization furrows in the dhc64Cm/z mutants (unpublished data), but during later cellularization, both are mislocalized basally along the full furrow length (Fig. 6 A, bracket; compare with WT, arrow). Baz and DE-Cad colocalize in many of the mislocalized puncta (Fig. 6 A, arrowhead). Thus, dynein functions to correctly position Baz and DE-Cad during cellularization; this could involve relatively direct dynein interactions or more indirect mechanisms. To our surprise, however, both Baz and DE-Cad become apically enriched as dhc64Cm/z mutants gastrulate (Fig. 6 D, arrows). Moreover, the dhc64Cm/z mutants produce large sheets of embryonic cuticle, indicative of relatively normal epithelial polarity (unpublished data; neighboring regions of missing cuticle are likely due to early syncytial defects). Thus, three mechanisms appear to position Baz during early development: an apical scaffold, dynein-mediated basal-to-apical transport and a third postcellularization activity.


The positioning and segregation of apical cues during epithelial polarity establishment in Drosophila.

Harris TJ, Peifer M - J. Cell Biol. (2005)

A specific role for dynein in positioning Baz and AJs. dhc64Cm/z mutants. (A–C) Cellularization. (A) Baz (red) and DE-Cad (green) mislocalize basally but often colocalize. WT, right. (B) aPKC (red) enriched apically. DE-Cad mislocalized basally (green). (C) PAR-6 (green) localizes as in WT. (D–G) Gastrulation. (D) Baz (red) and DE-Cad (green) are apical and colocalize. (E) aPKC apical. (F) PAR-6 apical. (G) Crb apical. (H and I) PMGIs. (H) aPKC (red) apical to DE-Cad (green) and Dlg (blue) segregates basally. (I) PAR-6 (green) above Baz (red) and Dlg (blue) basal. Bars, 5 μm.
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Related In: Results  -  Collection

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fig6: A specific role for dynein in positioning Baz and AJs. dhc64Cm/z mutants. (A–C) Cellularization. (A) Baz (red) and DE-Cad (green) mislocalize basally but often colocalize. WT, right. (B) aPKC (red) enriched apically. DE-Cad mislocalized basally (green). (C) PAR-6 (green) localizes as in WT. (D–G) Gastrulation. (D) Baz (red) and DE-Cad (green) are apical and colocalize. (E) aPKC apical. (F) PAR-6 apical. (G) Crb apical. (H and I) PMGIs. (H) aPKC (red) apical to DE-Cad (green) and Dlg (blue) segregates basally. (I) PAR-6 (green) above Baz (red) and Dlg (blue) basal. Bars, 5 μm.
Mentions: To directly test whether dynein functions in positioning Baz, we analyzed the Baz distribution in cellularizing dhc64Cm/z mutants. These mutants have defects in syncytial nuclear divisions (Robinson et al., 1999), but many undergo partial cellularization and initiate morphogenesis. In the mutants, centrosomes are positioned apically during cellularization (unpublished data), suggesting that basic MT polarity is retained. Baz and DE-Cad are recruited to early cellularization furrows in the dhc64Cm/z mutants (unpublished data), but during later cellularization, both are mislocalized basally along the full furrow length (Fig. 6 A, bracket; compare with WT, arrow). Baz and DE-Cad colocalize in many of the mislocalized puncta (Fig. 6 A, arrowhead). Thus, dynein functions to correctly position Baz and DE-Cad during cellularization; this could involve relatively direct dynein interactions or more indirect mechanisms. To our surprise, however, both Baz and DE-Cad become apically enriched as dhc64Cm/z mutants gastrulate (Fig. 6 D, arrows). Moreover, the dhc64Cm/z mutants produce large sheets of embryonic cuticle, indicative of relatively normal epithelial polarity (unpublished data; neighboring regions of missing cuticle are likely due to early syncytial defects). Thus, three mechanisms appear to position Baz during early development: an apical scaffold, dynein-mediated basal-to-apical transport and a third postcellularization activity.

Bottom Line: Adherens junctions (AJs) often direct this polarity, but we previously found that Bazooka (Baz) acts upstream of AJs as epithelial polarity is first established in Drosophila.Surprisingly, we found that Baz localizes to an apical domain below its typical binding partners atypical protein kinase C (aPKC) and partitioning defective (PAR)-6 as the Drosophila epithelium first forms.These results reveal key steps in the assembly of the apical domain in Drosophila.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA. tonyh@email.unc.edu

ABSTRACT
Cell polarity is critical for epithelial structure and function. Adherens junctions (AJs) often direct this polarity, but we previously found that Bazooka (Baz) acts upstream of AJs as epithelial polarity is first established in Drosophila. This prompted us to ask how Baz is positioned and how downstream polarity is elaborated. Surprisingly, we found that Baz localizes to an apical domain below its typical binding partners atypical protein kinase C (aPKC) and partitioning defective (PAR)-6 as the Drosophila epithelium first forms. In fact, Baz positioning is independent of aPKC and PAR-6 relying instead on cytoskeletal cues, including an apical scaffold and dynein-mediated basal-to-apical transport. AJ assembly is closely coupled to Baz positioning, whereas aPKC and PAR-6 are positioned separately. This forms a stratified apical domain with Baz and AJs localizing basal to aPKC and PAR-6, and we identify specific mechanisms that keep these proteins apart. These results reveal key steps in the assembly of the apical domain in Drosophila.

Show MeSH