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Drosophila melanogaster Cad99C, the orthologue of human Usher cadherin PCDH15, regulates the length of microvilli.

D'Alterio C, Tran DD, Yeung MW, Hwang MS, Li MA, Arana CJ, Mulligan VK, Kubesh M, Sharma P, Chase M, Tepass U, Godt D - J. Cell Biol. (2005)

Bottom Line: Loss of Cad99C function results in shortened and disorganized microvilli, whereas overexpression of Cad99C leads to a dramatic increase of microvillus length.Cad99C that lacks most of the cytoplasmic domain, including potential PDZ domain-binding sites, still promotes excessive microvillus outgrowth, suggesting that the amount of the extracellular domain determines microvillus length.This study reveals Cad99C as a critical regulator of microvillus length, the first example of a transmembrane protein that is involved in this process.

View Article: PubMed Central - PubMed

Affiliation: Department of Zoology, University of Toronto, Toronto, Ontario, Canada, M5S 3G5.

ABSTRACT
Actin-based protrusions can form prominent structures on the apical surface of epithelial cells, such as microvilli. Several cytoplasmic factors have been identified that control the dynamics of actin filaments in microvilli. However, it remains unclear whether the plasma membrane participates actively in microvillus formation. In this paper, we analyze the function of Drosophila melanogaster cadherin Cad99C in the microvilli of ovarian follicle cells. Cad99C contributes to eggshell formation and female fertility and is expressed in follicle cells, which produce the eggshells. Cad99C specifically localizes to apical microvilli. Loss of Cad99C function results in shortened and disorganized microvilli, whereas overexpression of Cad99C leads to a dramatic increase of microvillus length. Cad99C that lacks most of the cytoplasmic domain, including potential PDZ domain-binding sites, still promotes excessive microvillus outgrowth, suggesting that the amount of the extracellular domain determines microvillus length. This study reveals Cad99C as a critical regulator of microvillus length, the first example of a transmembrane protein that is involved in this process.

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Overexpression of Cad99C increases microvillus length. Transgenic proteins Cad99C-FL and Cad99CΔcyt::GFP were expressed in cell clones in the follicular epithelium using Act5c>CD2>Gal4. Cad99C is in red. (A–A”) Cells that express Cad99C-FL produced abnormally tall microvilli. A green arrowhead marks long microvilli projecting into wild-type territory. (B–B”) Expression of Cad99C-FL rescued the microvillus phenotype of a Cad99C21-5/21-6 mutant. (C–C”) Expression of Cad99CΔcyt::GFP induced lengthening of microvilli as seen with Cad99C (C, red) or GFP (C', green). (D–D”) Cad99CΔcyt::GFP expression rescued the microvilli in a Cad99C21-5/21-6 mutant follicle. Cad99C-FL (E and F) and Cad99CΔcyt::GFP (G) induce giant microvilli (green arrowheads) even in a Cad99C21-5/21-6 (E) or Cad99C21-8/21-6 (F and G) mutant background. (H) Structure of transgenic Cad99C-Fl and Cad99CΔcyt::GFP proteins. Confocal images are shown in A–G, and accompanying Nomarski images are shown in panels marked with ‘ or “. Yellow arrows mark clone boundaries in A–D. Bars, 10 μm.
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fig6: Overexpression of Cad99C increases microvillus length. Transgenic proteins Cad99C-FL and Cad99CΔcyt::GFP were expressed in cell clones in the follicular epithelium using Act5c>CD2>Gal4. Cad99C is in red. (A–A”) Cells that express Cad99C-FL produced abnormally tall microvilli. A green arrowhead marks long microvilli projecting into wild-type territory. (B–B”) Expression of Cad99C-FL rescued the microvillus phenotype of a Cad99C21-5/21-6 mutant. (C–C”) Expression of Cad99CΔcyt::GFP induced lengthening of microvilli as seen with Cad99C (C, red) or GFP (C', green). (D–D”) Cad99CΔcyt::GFP expression rescued the microvilli in a Cad99C21-5/21-6 mutant follicle. Cad99C-FL (E and F) and Cad99CΔcyt::GFP (G) induce giant microvilli (green arrowheads) even in a Cad99C21-5/21-6 (E) or Cad99C21-8/21-6 (F and G) mutant background. (H) Structure of transgenic Cad99C-Fl and Cad99CΔcyt::GFP proteins. Confocal images are shown in A–G, and accompanying Nomarski images are shown in panels marked with ‘ or “. Yellow arrows mark clone boundaries in A–D. Bars, 10 μm.

Mentions: To determine whether Cad99C is a limiting factor for microvillus outgrowth, we overexpressed full-length Cad99C (Cad99C-FL; Fig. 6 H) in follicle cells. Cad99C-FL expression induced a striking increase in the length of apical microvilli (Fig. 6 A). These long cellular protrusions, which sometimes exceeded the apical–basal length of the follicle cells, projected into the narrow space between follicle cells and oocyte (Fig. 6, A, E, and F). Very long extensions showed small knoblike dilatations. Interestingly, the vitelline bodies deposited between the overlong microvilli also appeared enlarged (Fig. 6 A'). Furthermore, expression of Cad99C-FL rescued microvillus formation in a Cad99C mutant background (Fig. 6 B). To investigate the importance of cytoplasmic interactions for Cad99C function, we expressed a transgenic protein (Cad99CΔcyt::GFP; Fig. 6 H) in which GFP replaces a large portion of the cytoplasmic tail, including the PDZ-binding sites. Strikingly, Cad99CΔcyt::GFP induced abnormally tall microvilli that fanned out from the apical surface and enlarged vitelline bodies similar to Cad99C-FL. This effect was observed in a wild-type (Fig. 6 C) and in a Cad99C mutant background (Fig. 6, D and G). These data suggest that Cad99C levels determine the length of microvilli and show that the majority of the cytoplasmic domain (256 out of 287 amino acids) can be deleted without affecting the potential of Cad99C to promote microvillus outgrowth.


Drosophila melanogaster Cad99C, the orthologue of human Usher cadherin PCDH15, regulates the length of microvilli.

D'Alterio C, Tran DD, Yeung MW, Hwang MS, Li MA, Arana CJ, Mulligan VK, Kubesh M, Sharma P, Chase M, Tepass U, Godt D - J. Cell Biol. (2005)

Overexpression of Cad99C increases microvillus length. Transgenic proteins Cad99C-FL and Cad99CΔcyt::GFP were expressed in cell clones in the follicular epithelium using Act5c>CD2>Gal4. Cad99C is in red. (A–A”) Cells that express Cad99C-FL produced abnormally tall microvilli. A green arrowhead marks long microvilli projecting into wild-type territory. (B–B”) Expression of Cad99C-FL rescued the microvillus phenotype of a Cad99C21-5/21-6 mutant. (C–C”) Expression of Cad99CΔcyt::GFP induced lengthening of microvilli as seen with Cad99C (C, red) or GFP (C', green). (D–D”) Cad99CΔcyt::GFP expression rescued the microvilli in a Cad99C21-5/21-6 mutant follicle. Cad99C-FL (E and F) and Cad99CΔcyt::GFP (G) induce giant microvilli (green arrowheads) even in a Cad99C21-5/21-6 (E) or Cad99C21-8/21-6 (F and G) mutant background. (H) Structure of transgenic Cad99C-Fl and Cad99CΔcyt::GFP proteins. Confocal images are shown in A–G, and accompanying Nomarski images are shown in panels marked with ‘ or “. Yellow arrows mark clone boundaries in A–D. Bars, 10 μm.
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fig6: Overexpression of Cad99C increases microvillus length. Transgenic proteins Cad99C-FL and Cad99CΔcyt::GFP were expressed in cell clones in the follicular epithelium using Act5c>CD2>Gal4. Cad99C is in red. (A–A”) Cells that express Cad99C-FL produced abnormally tall microvilli. A green arrowhead marks long microvilli projecting into wild-type territory. (B–B”) Expression of Cad99C-FL rescued the microvillus phenotype of a Cad99C21-5/21-6 mutant. (C–C”) Expression of Cad99CΔcyt::GFP induced lengthening of microvilli as seen with Cad99C (C, red) or GFP (C', green). (D–D”) Cad99CΔcyt::GFP expression rescued the microvilli in a Cad99C21-5/21-6 mutant follicle. Cad99C-FL (E and F) and Cad99CΔcyt::GFP (G) induce giant microvilli (green arrowheads) even in a Cad99C21-5/21-6 (E) or Cad99C21-8/21-6 (F and G) mutant background. (H) Structure of transgenic Cad99C-Fl and Cad99CΔcyt::GFP proteins. Confocal images are shown in A–G, and accompanying Nomarski images are shown in panels marked with ‘ or “. Yellow arrows mark clone boundaries in A–D. Bars, 10 μm.
Mentions: To determine whether Cad99C is a limiting factor for microvillus outgrowth, we overexpressed full-length Cad99C (Cad99C-FL; Fig. 6 H) in follicle cells. Cad99C-FL expression induced a striking increase in the length of apical microvilli (Fig. 6 A). These long cellular protrusions, which sometimes exceeded the apical–basal length of the follicle cells, projected into the narrow space between follicle cells and oocyte (Fig. 6, A, E, and F). Very long extensions showed small knoblike dilatations. Interestingly, the vitelline bodies deposited between the overlong microvilli also appeared enlarged (Fig. 6 A'). Furthermore, expression of Cad99C-FL rescued microvillus formation in a Cad99C mutant background (Fig. 6 B). To investigate the importance of cytoplasmic interactions for Cad99C function, we expressed a transgenic protein (Cad99CΔcyt::GFP; Fig. 6 H) in which GFP replaces a large portion of the cytoplasmic tail, including the PDZ-binding sites. Strikingly, Cad99CΔcyt::GFP induced abnormally tall microvilli that fanned out from the apical surface and enlarged vitelline bodies similar to Cad99C-FL. This effect was observed in a wild-type (Fig. 6 C) and in a Cad99C mutant background (Fig. 6, D and G). These data suggest that Cad99C levels determine the length of microvilli and show that the majority of the cytoplasmic domain (256 out of 287 amino acids) can be deleted without affecting the potential of Cad99C to promote microvillus outgrowth.

Bottom Line: Loss of Cad99C function results in shortened and disorganized microvilli, whereas overexpression of Cad99C leads to a dramatic increase of microvillus length.Cad99C that lacks most of the cytoplasmic domain, including potential PDZ domain-binding sites, still promotes excessive microvillus outgrowth, suggesting that the amount of the extracellular domain determines microvillus length.This study reveals Cad99C as a critical regulator of microvillus length, the first example of a transmembrane protein that is involved in this process.

View Article: PubMed Central - PubMed

Affiliation: Department of Zoology, University of Toronto, Toronto, Ontario, Canada, M5S 3G5.

ABSTRACT
Actin-based protrusions can form prominent structures on the apical surface of epithelial cells, such as microvilli. Several cytoplasmic factors have been identified that control the dynamics of actin filaments in microvilli. However, it remains unclear whether the plasma membrane participates actively in microvillus formation. In this paper, we analyze the function of Drosophila melanogaster cadherin Cad99C in the microvilli of ovarian follicle cells. Cad99C contributes to eggshell formation and female fertility and is expressed in follicle cells, which produce the eggshells. Cad99C specifically localizes to apical microvilli. Loss of Cad99C function results in shortened and disorganized microvilli, whereas overexpression of Cad99C leads to a dramatic increase of microvillus length. Cad99C that lacks most of the cytoplasmic domain, including potential PDZ domain-binding sites, still promotes excessive microvillus outgrowth, suggesting that the amount of the extracellular domain determines microvillus length. This study reveals Cad99C as a critical regulator of microvillus length, the first example of a transmembrane protein that is involved in this process.

Show MeSH
Related in: MedlinePlus