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On the role of the MAGUK proteins encoded by Drosophila varicose during embryonic and postembryonic development.

Bachmann A, Draga M, Grawe F, Knust E - BMC Dev. Biol. (2008)

Bottom Line: Their capacity to serve as platforms for organising larger protein assemblies results from the presence of several protein-protein interaction domains.Postembryonic reduction of varicose function by expressing double-stranded RNA affects pattern formation and morphogenesis of the wing and the development of normal-shaped and -sized eyes.Expression of two Varicose isoforms in embryonic epithelia and imaginal discs suggests that the composition of Varicose-mediated protein scaffolds at septate junctions is dynamic, which may have important implications for the modulation of their function.

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Affiliation: Institut für Genetik, Heinrich-Heine-Universität Düsseldorf, Universitätsstr. 1, 40225 Düsseldorf, Germany. bachmana@uni-duesseldorf.de

ABSTRACT

Background: Membrane-associated guanylate kinases (MAGUKs) form a family of scaffolding proteins, which are often associated with cellular junctions, such as the vertebrate tight junction, the Drosophila septate junction or the neuromuscular junction. Their capacity to serve as platforms for organising larger protein assemblies results from the presence of several protein-protein interaction domains. They often appear in different variants suggesting that they also mediate dynamic changes in the composition of the complexes.

Results: Here we show by electron microscopic analysis that Drosophila embryos lacking varicose function fail to develop septate junctions in the tracheae and the epidermis. In the embryo and in imaginal discs varicose expresses two protein isoforms, which belong to the MAGUK family. The two isoforms can be distinguished by the presence or absence of two L27 domains and are differentially affected in different varicose alleles. While the short isoform is essential for viability, the long isoform seems to have a supportive function. Varicose proteins co-localise with Neurexin IV in pleated septate junctions and are necessary, but not sufficient for its recruitment. The two proteins interact in vitro by the PDZ domain of Varicose and the four C-terminal amino acids of Neurexin IV. Postembryonic reduction of varicose function by expressing double-stranded RNA affects pattern formation and morphogenesis of the wing and the development of normal-shaped and -sized eyes.

Conclusion: Expression of two Varicose isoforms in embryonic epithelia and imaginal discs suggests that the composition of Varicose-mediated protein scaffolds at septate junctions is dynamic, which may have important implications for the modulation of their function.

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EM analysis reveals defective septate junctions in vari mutants. Epidermis (A-D, I) and tracheae (E-H) of stage 16 embryos. (A, E): variMD109/CyO-twi-GFP; (B, F): variMD109; (C, G): vari03953b/CyO-twi-GFP; (D, H): vari03953b; (I): wild-type. Adherens junctions (black arrowheads) and developing septate junctions (white arrows) can be distinguished in wild-type embryos. The inset in E shows the regularly aligned septa of pleated septate junctions between adjacent cells. Septate junctions are absent in homozygous variMD109 (B, F) and vari03953b (D, H) mutant embryos, while adherens junctions are well developed (black arrowheads).
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Figure 4: EM analysis reveals defective septate junctions in vari mutants. Epidermis (A-D, I) and tracheae (E-H) of stage 16 embryos. (A, E): variMD109/CyO-twi-GFP; (B, F): variMD109; (C, G): vari03953b/CyO-twi-GFP; (D, H): vari03953b; (I): wild-type. Adherens junctions (black arrowheads) and developing septate junctions (white arrows) can be distinguished in wild-type embryos. The inset in E shows the regularly aligned septa of pleated septate junctions between adjacent cells. Septate junctions are absent in homozygous variMD109 (B, F) and vari03953b (D, H) mutant embryos, while adherens junctions are well developed (black arrowheads).

Mentions: Based on the failure to properly localise the septate-junction-associated proteins Neurexin IV and Coracle (Cora) in vari mutant embryos (Fig. 3 and [13]) Wu et al. suggested that these embryos fail to properly establish and/or maintain the septate junctions. In order to prove this, we carried out electron microscopic analysis of wild-type (or heterozygous) and vari mutant embryos. In wild-type embryos, pleated septate junctions are clearly distinguishable from stage 16 onwards [9] by the presence of septa, which span the space between the lateral membranes of neighbouring cells, both in the epidermis (Fig. 4A,C,I) and the tracheae (Fig. 4E,G). Homozygous mutant variMD109 embryos of the same stage lacked septate junctions in these tissues (Fig. 4B,F). Similarly, septate junctions were not detected in vari03953b homozygous mutant embryos (Fig. 4D,H). The ZA was not affected in epithelia of the mutant embryos and still formed a continuous apical belt. This result finally proves that vari has an essential function in the formation of the pleated septate junctions during embryonic development.


On the role of the MAGUK proteins encoded by Drosophila varicose during embryonic and postembryonic development.

Bachmann A, Draga M, Grawe F, Knust E - BMC Dev. Biol. (2008)

EM analysis reveals defective septate junctions in vari mutants. Epidermis (A-D, I) and tracheae (E-H) of stage 16 embryos. (A, E): variMD109/CyO-twi-GFP; (B, F): variMD109; (C, G): vari03953b/CyO-twi-GFP; (D, H): vari03953b; (I): wild-type. Adherens junctions (black arrowheads) and developing septate junctions (white arrows) can be distinguished in wild-type embryos. The inset in E shows the regularly aligned septa of pleated septate junctions between adjacent cells. Septate junctions are absent in homozygous variMD109 (B, F) and vari03953b (D, H) mutant embryos, while adherens junctions are well developed (black arrowheads).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 4: EM analysis reveals defective septate junctions in vari mutants. Epidermis (A-D, I) and tracheae (E-H) of stage 16 embryos. (A, E): variMD109/CyO-twi-GFP; (B, F): variMD109; (C, G): vari03953b/CyO-twi-GFP; (D, H): vari03953b; (I): wild-type. Adherens junctions (black arrowheads) and developing septate junctions (white arrows) can be distinguished in wild-type embryos. The inset in E shows the regularly aligned septa of pleated septate junctions between adjacent cells. Septate junctions are absent in homozygous variMD109 (B, F) and vari03953b (D, H) mutant embryos, while adherens junctions are well developed (black arrowheads).
Mentions: Based on the failure to properly localise the septate-junction-associated proteins Neurexin IV and Coracle (Cora) in vari mutant embryos (Fig. 3 and [13]) Wu et al. suggested that these embryos fail to properly establish and/or maintain the septate junctions. In order to prove this, we carried out electron microscopic analysis of wild-type (or heterozygous) and vari mutant embryos. In wild-type embryos, pleated septate junctions are clearly distinguishable from stage 16 onwards [9] by the presence of septa, which span the space between the lateral membranes of neighbouring cells, both in the epidermis (Fig. 4A,C,I) and the tracheae (Fig. 4E,G). Homozygous mutant variMD109 embryos of the same stage lacked septate junctions in these tissues (Fig. 4B,F). Similarly, septate junctions were not detected in vari03953b homozygous mutant embryos (Fig. 4D,H). The ZA was not affected in epithelia of the mutant embryos and still formed a continuous apical belt. This result finally proves that vari has an essential function in the formation of the pleated septate junctions during embryonic development.

Bottom Line: Their capacity to serve as platforms for organising larger protein assemblies results from the presence of several protein-protein interaction domains.Postembryonic reduction of varicose function by expressing double-stranded RNA affects pattern formation and morphogenesis of the wing and the development of normal-shaped and -sized eyes.Expression of two Varicose isoforms in embryonic epithelia and imaginal discs suggests that the composition of Varicose-mediated protein scaffolds at septate junctions is dynamic, which may have important implications for the modulation of their function.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institut für Genetik, Heinrich-Heine-Universität Düsseldorf, Universitätsstr. 1, 40225 Düsseldorf, Germany. bachmana@uni-duesseldorf.de

ABSTRACT

Background: Membrane-associated guanylate kinases (MAGUKs) form a family of scaffolding proteins, which are often associated with cellular junctions, such as the vertebrate tight junction, the Drosophila septate junction or the neuromuscular junction. Their capacity to serve as platforms for organising larger protein assemblies results from the presence of several protein-protein interaction domains. They often appear in different variants suggesting that they also mediate dynamic changes in the composition of the complexes.

Results: Here we show by electron microscopic analysis that Drosophila embryos lacking varicose function fail to develop septate junctions in the tracheae and the epidermis. In the embryo and in imaginal discs varicose expresses two protein isoforms, which belong to the MAGUK family. The two isoforms can be distinguished by the presence or absence of two L27 domains and are differentially affected in different varicose alleles. While the short isoform is essential for viability, the long isoform seems to have a supportive function. Varicose proteins co-localise with Neurexin IV in pleated septate junctions and are necessary, but not sufficient for its recruitment. The two proteins interact in vitro by the PDZ domain of Varicose and the four C-terminal amino acids of Neurexin IV. Postembryonic reduction of varicose function by expressing double-stranded RNA affects pattern formation and morphogenesis of the wing and the development of normal-shaped and -sized eyes.

Conclusion: Expression of two Varicose isoforms in embryonic epithelia and imaginal discs suggests that the composition of Varicose-mediated protein scaffolds at septate junctions is dynamic, which may have important implications for the modulation of their function.

Show MeSH
Related in: MedlinePlus