InteractiveFly: GeneBrief

Exo84: Biological Overview | References

Gene name - Exo84

Synonyms -

Cytological map position- 96F10-96F10

Function - signaling

Keywords - exocyst complex subunit, apical polarity, polarized exocytosis, vesicular transport vesicles

Symbol - Exo84

FlyBase ID: FBgn0266668

Genetic map position - 3R: 21,875,452..21,877,957 [-]

Classification - Exocyst complex 84-kDa subunit Pleckstrin Homology (PH) domain

Cellular location - cytoplasmic

NCBI links: Precomputed BLAST | EntrezGene

The polarized architecture of epithelial tissues involves a dynamic balance between apical and basolateral membrane domains. This study shows that epithelial polarity in the Drosophila embryo requires the exocyst complex subunit homolog Exo84. Exo84 activity is essential for the apical localization of the Crumbs transmembrane protein, a key determinant of epithelial apical identity. Adherens junction proteins become mislocalized at the cell surface in Exo84 mutants in a pattern characteristic of defects in apical, but not basolateral, components. Loss of Crumbs from the cell surface precedes the disruption of Bazooka and Armadillo localization in Exo84 mutants. Moreover, Exo84 mutants display defects in apical cuticle secretion that are similar to crumbs mutants and are suppressed by a reduction in the basolateral proteins Dlg and Lgl. In Exo84 mutants at advanced stages of epithelial degeneration, apical and adherens junction proteins accumulate in an expanded recycling endosome compartment. These results suggest that epithelial polarity in the Drosophila embryo is actively maintained by exocyst-dependent apical localization of the Crumbs transmembrane protein (Blankenship, 2007).

Epithelial cells in the Drosophila embryo generate molecularly distinct apical and basolateral surfaces that provide structural integrity to the developing embryo. Specialized cell surface domains are separated by intercellular adherens junctions that initiate as diffuse apicolateral accumulations and subsequently coalesce to form a discrete apical band called the zonula adherens. The spatial organization of mature adherens junctions is actively maintained by input from both apical and basolateral proteins. The Crumbs EGF-repeat transmembrane protein and its cytoplasmic binding partners Stardust and PATJ localize to the apical cell surface and are required for epithelial structure and adherens junction morphology. In addition, overexpression of Crumbs leads to a selective expansion of the apical cell surface, demonstrating that Crumbs is necessary and sufficient for apical identity. The localization of mature adherens junctions also requires the basolateral PDZ-domain proteins Discs large (Dlg) and Scribble (Scrib) and the WD40-domain protein Lethal giant larvae (Lgl) Epithelial defects caused by disruption of apical Crumbs activity can be rescued by a simultaneous reduction in the activity of basolateral proteins, indicating that apical and basolateral domains function in opposition to maintain epithelial polarity (Blankenship, 2007 and references therein).

Misregulation of Crumbs activity can have severe effects on cell and tissue function and is associated with human retinal diseases. Multiple mechanisms contribute to Crumbs localization, stability and activity to precisely control its function. The basolateral proteins Dlg, Lgl and Scrib oppose Crumbs activity and restrict its localization in the Drosophila embryo, and the Yurt FERM-domain protein associates with the Crumbs cytoplasmic domain and negatively regulates Crumbs activity at the apicolateral cell surface. Endocytosis of Crumbs protein is also required for tissue morphology; mutations in the Avalanche syntaxin or the Rab5 GTPase lead to Crumbs accumulation and wing imaginal disc overgrowth (Lu, 2005). In addition, a complex containing the Rich1 Cdc42 GAP protein and the angiomotin scaffolding protein associates with cytoplasmic binding partners of Crumbs and provides a potential link between the Crumbs complex and the endocytic machinery (Wells, 2006). However, the mechanisms that govern the delivery of Crumbs protein to the cell surface are not known (Blankenship, 2007).

The targeting of transmembrane proteins to specific destinations at the cell surface is a widely used mechanism for establishing cell polarity. The spatial specificity of vesicle trafficking is thought to occur at a late step in this process through the tethering of exocytic vesicles at defined membrane sites by the eight-subunit exocyst (or Sec6/8) complex (Lipschutz, 2002; Whyte, 2002). Exocyst components were originally identified based on their role in polarized secretion in Saccharomyces cerevisiae and were subsequently shown to form a complex that is highly conserved from yeast to mammals. In multicellular organisms, exocyst components are required for multiple developmental processes including epithelial polarity, membrane integrity, photoreceptor morphogenesis, cell fate determination and synapse formation. These diverse functions demonstrate that polarized exocytosis is a fundamental mechanism for regulating cell morphology (Blankenship, 2007).

This study provided evidence that the Drosophila homolog of the Exo84 exocyst complex subunit is essential for epithelial polarity and apical protein localization in the Drosophila embryo. In Exo84 mutants, adherens junction proteins become mislocalized along the apical-basal axis in a manner reminiscent of cells lacking the Crumbs apical determinant. Loss of Crumbs from the apical surface is the earliest defect detected in Exo84 mutants. Exo84 mutants at advanced stages of epithelial degeneration display defects in trafficking apical and junctional proteins from the recycling endosome to the cell surface. These results demonstrate that the Drosophila homolog of the exocyst complex subunit Exo84 plays an essential role in epithelial polarity by regulating the localization of the Crumbs apical determinant (Blankenship, 2007).

It is concluded that epithelial polarity in the Drosophila embryo is actively maintained by the Exo84-dependent localization of the Crumbs transmembrane protein to the apical surface. Exo84 mutants display an aberrant distribution of junctional proteins that resembles the phenotype of crumbs mutants, and depletion of Crumbs from the apical surface is the earliest defect detected in Exo84 mutants. In addition, the onset of epithelial disruption at stage 9 in Exo84 mutants is comparable with the timing of the crumbs mutant defects, and the Crumbs protein still aggregates in Exo84 embryos with greatly reduced E-cadherin. Exo84 is likely to function as part of the exocyst complex in the Drosophila embryo, in light of the genetic interactions observed between Exo84 and the Sec5 and Sec6 exocyst subunits and the common defects in recycling endosome morphology caused by exocyst disruption in multiple cellular contexts (Jafar-Nejad, 2005; Langevin, 2005). These results suggest a role for exocyst-dependent membrane trafficking in the maintenance of apical epithelial identity in the Drosophila embryo (Blankenship, 2007).

In contrast to the relatively specific mislocalization of Crumbs in stage 9 Exo84 mutant embryos, by late stage 10 these embryos display defects in the delivery of multiple proteins to the cell surface. Epithelial polarity and the distribution of apical and junctional proteins are established correctly in Exo84 mutants, either because these processes occur independently of Exo84 or because of residual Exo84 activity in this mutant background. The earliest defect observed in Exo84 mutants is a loss of Crumbs from the apical surface during epithelial maturation. As a likely consequence of the loss of cell-surface Crumbs localization, adherens junction proteins become mislocalized to varying positions along the basolateral cell membrane. Mutant embryos at later stages display a cytoplasmic accumulation of apical and adherens junction proteins in an expanded Rab11 recycling endosome compartment, consistent with a defect in vesicular transport to the cell surface. The failure to deliver junctional proteins to the cell surface is unlikely to result from a defect in Crumbs localization, because the cytoplasmic accumulation of junctional proteins does not occur in crumbs mutants. These results indicate that disruption of exocyst-dependent membrane trafficking ultimately results in the failure to deliver both apical and junctional proteins from the recycling endosome to the cell surface. The mislocalization of apical and junctional proteins in Exo84 mutant embryos is associated with a loss of columnar morphology, demonstrating that Exo84 activity is essential for epithelial organization (Blankenship, 2007).

A precise balance between apical and basolateral determination is essential for epithelial integrity and the placement of the zonula adherens in the Drosophila embryo. This balance is actively maintained by Exo84-dependent localization of the Crumbs transmembrane protein to the apical cell surface. Loss of apical or basolateral identity leads to distinct patterns of junctional protein distribution, suggesting that the apical and basal limits of the zonula adherens are defined by different mechanisms. In crumbs mutants, DE-cadherin and Armadillo are restricted to focused puncta at varying locations at the cell surface. By contrast, in embryos defective for the basolateral proteins Dlg and Lgl, junctional proteins are dispersed along the plasma membrane rather than aggregating at a single site. A basolateral expansion of the apical Crumbs domain has also been reported in dlg and lgl mutants. These results suggest that basolateral proteins create a nonpermissive barrier to adherens junction expansion, whereas apical proteins may play a positive role in recruiting or stabilizing junctions at the apical cell surface. Consistent with this possibility, the apical Crumbs domain is closely apposed to the zonula adherens, and it was found that Bazooka and Armadillo colocalize at the cell surface and in the cytoplasm of Exo84 mutant embryos. Exocyst-dependent trafficking of Crumbs to the apical surface may reinforce the apical epithelial domain and stabilize the apicolateral localization of the zonula adherens (Blankenship, 2007).

The recycling endosome is the primary vesicular compartment affected in embryos mutant for the exocyst subunit homolog Exo84, while Golgi, early endosomal and late endosomal compartments remain largely intact. Exocyst proteins are required for recycling endosome morphology in several epithelial and sensory cell types (Jafar-Nejad, 2005; Langevin, 2005) and the Rab11 recycling endosome protein can associate directly with the exocyst subunits Sec5 and Sec15. It was found that Rab11 vesicles in maturing embryonic epithelia are enriched in the apical cytoplasm, where they preferentially accumulate in the plane of the adherens junctions. Conversely, a basal expansion of recycling endosomes during cellularization correlates with a basal bias in membrane addition. These results suggest that there is a spatial correlation between the sites of recycling endosome accumulation and the surface destinations of proteins trafficked through recycling endosomes. The redistribution of the recycling endosome compartment to the apical cytoplasm accompanies the transition from basolateral to apical membrane insertion and may reflect the onset of a critical requirement for Crumbs activity during epithelial maturation (Blankenship, 2007).

The requirement for Exo84 in apical protein localization in the Drosophila embryo is distinct from exocyst functions in other epithelia, in which exocyst components are required for the localization of basolateral or junctional proteins. The results indicate that Exo84 is also required for delivery of DE-cadherin to the cell surface in the embryo, consistent with the demonstrated roles for Sec5, Sec6 and Sec15 in DE-cadherin trafficking in the pupal epithelium (Langevin, 2005). However, although the mislocalization of the apical Crumbs protein is a primary defect of Exo84 mutant embryos, exocyst mutations do not appreciably affect Crumbs localization in pupal epithelial and photoreceptor cells (Beronja, 2005; Langevin, 2005). These results are consistent with a model in which distinct cargo proteins are trafficked by the exocyst complex in different cellular contexts. Alternatively, DE-cadherin and Crumbs may be delivered to the cell surface in an exocyst-dependent fashion in multiple cell types, but undergo different rates of turnover. For example, Crumbs may be dynamically trafficked in the embryo but stably maintained at the surface of pupal epithelial cells. Differential effects on specific target proteins are not atypical of exocyst function, because loss of Sec6 activity in Drosophila photoreceptor cells disrupts the localization of the rhabdomere proteins Chaoptin and Rhodopsin1, whereas the apical localization of Crumbs and DE-cadherin occurs normally (Beronja, 2005). The Drosophila embryonic epithelium undergoes pronounced changes in structure and organization during development that rely on a balance between apical and basolateral surface domains. A requirement for exocyst-dependent Crumbs trafficking during this process may facilitate the dynamic remodeling of epithelial polarity during morphogenesis (Blankenship, 2007).


Search PubMed for articles about Drosophila Exo84

Beronja, S., Laprise, P., Papoulas, O., Pellikka, M., Sisson, J. and Tepass, U. (2005). Essential function of Drosophila Sec6 in apical exocytosis of epithelial photoreceptor cells. J. Cell Biol. 169(4): 635-46. PubMed ID: 15897260

Blankenship, J. T., Fuller, M. T. and Zallen, J. A. (2007). The Drosophila homolog of the Exo84 exocyst subunit promotes apical epithelial identity. J. Cell Sci. 120: 3099-3110. PubMed ID: 17698923

Jafar-Nejad, H., Andrews, H. K., Acar, M., Bayat, V., Wirtz-Peitz, F., Mehta, S. Q., Knoblich, J. A. and Bellen, H. J. (2005). Sec15, a component of the exocyst, promotes notch signaling during the asymmetric division of Drosophila sensory organ precursors. Dev. Cell 9(3): 351-63. PubMed ID: 16137928

Langevin, J., et al. (2005). Drosophila exocyst components Sec5, Sec6, and Sec15 regulate DE-Cadherin trafficking from recycling endosomes to the plasma membrane. Dev. Cell 9(3): 355-76. PubMed ID: 16224820

Lipschutz, J. H. and Mostov, K. E. (2002). Exocytosis: the many masters of the exocyst. Curr. Biol. 12: 212-214. PubMed ID: 9136678

Lu, H. and Bilder, D. (2005). Endocytic control of epithelial polarity and proliferation in Drosophila. Nat. Cell Biol. 7: 1232-1239. PubMed ID: 16258546

Wells, C. D., Fawcett, J. P., Traweger, A., Yamanaka, Y., Goudreault, M., Elder, K., Kulkarni, S., Gish, G., Virag, C., Lim, C., et al. (2006). Rich1/Amot complex regulates the Cdc42 GTPase and apical-polarity proteins in epithelial cells. Cell 125: 535-548. PubMed ID: 16678097

Whyte, J. R. and Munro, S. (2002). Vesicle tethering complexes in membrane traffic. J. Cell Sci. 115: 2627-2637. PubMed ID: 12077354

Biological Overview

date revised: 30 December 2007

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