Patricia S. Connelly

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

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 μm in length); modules elongate to ∼3 μm 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.

Microvilli appear to represent the first step in actin bundle formation in Drosophila bristles

During bristle development the emerging bristle shaft, socket cell, and the apical surface of thoracic epithelial cells form tiny protuberances or pimples that contain electron-dense material located on the cytoplasmic surface of the pimple tip. In a few cases short actin filaments extend from this material into the cortical cytoplasm. When cultured in the presence of jasplakinolide, an agent that prevents filament disassembly, pimples elongate to form microvilli containing a core of crosslinked filaments. Emerging-bristle mutants delay cortical bundle formation and are aggregated by forked protein crossbridges. Using these mutants and enhancing core bundle formation with jasplakinolide we found that microvillar formation represents the first stage in the morphogenesis of much larger actin bundles in Drosophila bristle shaft cells. Evidence is presented showing that socket cells do not contain forked protein crossbridges, a fact that may explain why cortical bundles only appear in bristle shaft cells. Furthermore, as pimples and microvilli form in the absence of both forked and fascin crossbridges, we also conclude that neither of these crossbridges account for core bundle formation in microvilli, but there must exist a third, as yet unidentified crossbridge in this system. Immunocytochemisty suggested that this new crossbridge is not Drosophila villin. Finally, ultrastructural comparisons suggest that microspikes and microvilli form very differently.