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Long continuous actin bundles in Drosophila bristles are constructed by overlapping short filaments.

Guild GM, Connelly PS, Ruggiero L, Vranich KA, Tilney LG - J. Cell Biol. (2003)

Bottom Line: These long bundles are built from much shorter modules that graft together.Thus, bundle morphogenesis has several components: module formation, elongation, grafting, and bundle smoothing.These actin bundles are much like a rope or cable, made by overlapping elements that run a small fraction of the overall length, and stiffened by cross-linking.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Pennsylvania, Philadelphia, PA 19104-6018, USA. gguild@sas.upenn.edu

ABSTRACT
The actin bundles essential for Drosophila bristle elongation are hundreds of microns long and composed of cross-linked unipolar filaments. These long bundles are built from much shorter modules that graft together. Using both confocal and electron microscopy, we demonstrate that newly synthesized modules are short (1-2 microm in length); modules elongate to approximately 3 microm by growing over the surface of longitudinally adjacent modules to form a graft; the grafted regions are initially secured by the forked protein cross-bridge and later by the fascin cross-bridge; actin bundles are smoothed by filament addition and appear continuous and without swellings; and in the absence of grafting, dramatic alterations in cell shape occur that substitutes cell width expansion for elongation. Thus, bundle morphogenesis has several components: module formation, elongation, grafting, and bundle smoothing. These actin bundles are much like a rope or cable, made by overlapping elements that run a small fraction of the overall length, and stiffened by cross-linking.

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Actin modules overlap during grafting. (a and b) Confocal images of growing bristle tips from 44-h wild-type macrochaetes stained with rhodamine phalloidin showing overlapping modules (arrowheads). (c–f) Confocal micrographs of a wild-type 36-h pupal bristle stained with antiforked antibody (green) and phalloidin (red). A macrochaete showing thin actin bundles (d) containing the forked cross-bridge (c). Periodic “knuckles” representing module overlap can be seen along the bundles using the antiforked antibody. Many of these knuckles are in transverse register (e.g., arrowheads). (e and f) A higher magnification view of four actin-forked knuckles shown in c and d between the arrowheads. As expected, sprouting bristles that lack forked protein (forked36a mutant) were not stained with the forked antibody but were stained with phalloidin (not depicted). Bars: (a and b) 2 μm; (c and d) 5 μm.
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fig4: Actin modules overlap during grafting. (a and b) Confocal images of growing bristle tips from 44-h wild-type macrochaetes stained with rhodamine phalloidin showing overlapping modules (arrowheads). (c–f) Confocal micrographs of a wild-type 36-h pupal bristle stained with antiforked antibody (green) and phalloidin (red). A macrochaete showing thin actin bundles (d) containing the forked cross-bridge (c). Periodic “knuckles” representing module overlap can be seen along the bundles using the antiforked antibody. Many of these knuckles are in transverse register (e.g., arrowheads). (e and f) A higher magnification view of four actin-forked knuckles shown in c and d between the arrowheads. As expected, sprouting bristles that lack forked protein (forked36a mutant) were not stained with the forked antibody but were stained with phalloidin (not depicted). Bars: (a and b) 2 μm; (c and d) 5 μm.

Mentions: We looked for evidence for module–module overlap before bundle smoothing by examining bristle tips stained with fluorescent phalloidin. By acquiring thin confocal sections we could image modules in profile that had elongated over one another before grafting (Fig. 4, a and b).


Long continuous actin bundles in Drosophila bristles are constructed by overlapping short filaments.

Guild GM, Connelly PS, Ruggiero L, Vranich KA, Tilney LG - J. Cell Biol. (2003)

Actin modules overlap during grafting. (a and b) Confocal images of growing bristle tips from 44-h wild-type macrochaetes stained with rhodamine phalloidin showing overlapping modules (arrowheads). (c–f) Confocal micrographs of a wild-type 36-h pupal bristle stained with antiforked antibody (green) and phalloidin (red). A macrochaete showing thin actin bundles (d) containing the forked cross-bridge (c). Periodic “knuckles” representing module overlap can be seen along the bundles using the antiforked antibody. Many of these knuckles are in transverse register (e.g., arrowheads). (e and f) A higher magnification view of four actin-forked knuckles shown in c and d between the arrowheads. As expected, sprouting bristles that lack forked protein (forked36a mutant) were not stained with the forked antibody but were stained with phalloidin (not depicted). Bars: (a and b) 2 μm; (c and d) 5 μm.
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Related In: Results  -  Collection

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fig4: Actin modules overlap during grafting. (a and b) Confocal images of growing bristle tips from 44-h wild-type macrochaetes stained with rhodamine phalloidin showing overlapping modules (arrowheads). (c–f) Confocal micrographs of a wild-type 36-h pupal bristle stained with antiforked antibody (green) and phalloidin (red). A macrochaete showing thin actin bundles (d) containing the forked cross-bridge (c). Periodic “knuckles” representing module overlap can be seen along the bundles using the antiforked antibody. Many of these knuckles are in transverse register (e.g., arrowheads). (e and f) A higher magnification view of four actin-forked knuckles shown in c and d between the arrowheads. As expected, sprouting bristles that lack forked protein (forked36a mutant) were not stained with the forked antibody but were stained with phalloidin (not depicted). Bars: (a and b) 2 μm; (c and d) 5 μm.
Mentions: We looked for evidence for module–module overlap before bundle smoothing by examining bristle tips stained with fluorescent phalloidin. By acquiring thin confocal sections we could image modules in profile that had elongated over one another before grafting (Fig. 4, a and b).

Bottom Line: These long bundles are built from much shorter modules that graft together.Thus, bundle morphogenesis has several components: module formation, elongation, grafting, and bundle smoothing.These actin bundles are much like a rope or cable, made by overlapping elements that run a small fraction of the overall length, and stiffened by cross-linking.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Pennsylvania, Philadelphia, PA 19104-6018, USA. gguild@sas.upenn.edu

ABSTRACT
The actin bundles essential for Drosophila bristle elongation are hundreds of microns long and composed of cross-linked unipolar filaments. These long bundles are built from much shorter modules that graft together. Using both confocal and electron microscopy, we demonstrate that newly synthesized modules are short (1-2 microm in length); modules elongate to approximately 3 microm by growing over the surface of longitudinally adjacent modules to form a graft; the grafted regions are initially secured by the forked protein cross-bridge and later by the fascin cross-bridge; actin bundles are smoothed by filament addition and appear continuous and without swellings; and in the absence of grafting, dramatic alterations in cell shape occur that substitutes cell width expansion for elongation. Thus, bundle morphogenesis has several components: module formation, elongation, grafting, and bundle smoothing. These actin bundles are much like a rope or cable, made by overlapping elements that run a small fraction of the overall length, and stiffened by cross-linking.

Show MeSH
Related in: MedlinePlus