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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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Baz positioning involves basal-to-apical transport and dynein. (A) BazGFP puncta moving apically at late cellularization (4 s intervals; Video S3). Velocity distribution in histogram (right). (B) Basal BazGFP puncta (green) overlap with DIC (red; arrows). Insets show overlap after CD treatment. (C) Cuticles. Most baz zygotic mutants have one cuticle hole (left, arrow). dhc64C mutations enhance the baz phenotype, producing larger holes (right, arrows), and overall loss of cuticle (right, outlined). Quantification below. Classes with >20% of dead embryos in bold. Bars, 5 μm.
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fig5: Baz positioning involves basal-to-apical transport and dynein. (A) BazGFP puncta moving apically at late cellularization (4 s intervals; Video S3). Velocity distribution in histogram (right). (B) Basal BazGFP puncta (green) overlap with DIC (red; arrows). Insets show overlap after CD treatment. (C) Cuticles. Most baz zygotic mutants have one cuticle hole (left, arrow). dhc64C mutations enhance the baz phenotype, producing larger holes (right, arrows), and overall loss of cuticle (right, outlined). Quantification below. Classes with >20% of dead embryos in bold. Bars, 5 μm.

Mentions: A later Baz-positioning mechanism was revealed when we analyzed the ectopic BazGFP puncta at the end of cellularization. In 14/16 embryos with ectopic BazGFP puncta during cellularization (Fig. 4 B, 0:14 and 0:23, arrowheads), the basal BazGFP was cleared by gastrulation onset, restoring nearly normal localization (Fig. 4 B, 0:31, arrow). Moreover, 70% of BazGFP embryos complete embryogenesis and the 30% that die have head holes but an otherwise normal embryonic cuticle, indicative of normal epithelial polarity (unpublished data). By examining cellularizing embryos, we observed basal BazGFP puncta undergoing basal-to-apical translocations as the furrows pass the base of the nucleus (Fig. 5 A, arrows; Video S3 available at http://www.jcb.org/cgi/content/full/jcb.200505127/DC1). Particles move at 183 ± 60 nm/s (n = 42; Fig. 5 A), progressively clearing ectopic BazGFP from basal regions. Thus, endogenous Baz positioning may also involve basal-to-apical transport.


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

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

Baz positioning involves basal-to-apical transport and dynein. (A) BazGFP puncta moving apically at late cellularization (4 s intervals; Video S3). Velocity distribution in histogram (right). (B) Basal BazGFP puncta (green) overlap with DIC (red; arrows). Insets show overlap after CD treatment. (C) Cuticles. Most baz zygotic mutants have one cuticle hole (left, arrow). dhc64C mutations enhance the baz phenotype, producing larger holes (right, arrows), and overall loss of cuticle (right, outlined). Quantification below. Classes with >20% of dead embryos in bold. Bars, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171335&req=5

fig5: Baz positioning involves basal-to-apical transport and dynein. (A) BazGFP puncta moving apically at late cellularization (4 s intervals; Video S3). Velocity distribution in histogram (right). (B) Basal BazGFP puncta (green) overlap with DIC (red; arrows). Insets show overlap after CD treatment. (C) Cuticles. Most baz zygotic mutants have one cuticle hole (left, arrow). dhc64C mutations enhance the baz phenotype, producing larger holes (right, arrows), and overall loss of cuticle (right, outlined). Quantification below. Classes with >20% of dead embryos in bold. Bars, 5 μm.
Mentions: A later Baz-positioning mechanism was revealed when we analyzed the ectopic BazGFP puncta at the end of cellularization. In 14/16 embryos with ectopic BazGFP puncta during cellularization (Fig. 4 B, 0:14 and 0:23, arrowheads), the basal BazGFP was cleared by gastrulation onset, restoring nearly normal localization (Fig. 4 B, 0:31, arrow). Moreover, 70% of BazGFP embryos complete embryogenesis and the 30% that die have head holes but an otherwise normal embryonic cuticle, indicative of normal epithelial polarity (unpublished data). By examining cellularizing embryos, we observed basal BazGFP puncta undergoing basal-to-apical translocations as the furrows pass the base of the nucleus (Fig. 5 A, arrows; Video S3 available at http://www.jcb.org/cgi/content/full/jcb.200505127/DC1). Particles move at 183 ± 60 nm/s (n = 42; Fig. 5 A), progressively clearing ectopic BazGFP from basal regions. Thus, endogenous Baz positioning may also involve basal-to-apical transport.

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