A high resolution view of the fly actin cytoskeleton lacking a functional WAVE complex

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Abstract

The development of multicellular organisms involves a series of morphogenetic processes coordinating a highly dynamic and organized interplay between cells and their environment. Thus, the generation of forces that drive cellular and intracellular movements is prerequisite to shape single cells into tissues and organs. The actin cytoskeleton represents a highly dynamic filamentous system providing cell structure and mechanical forces to drive membrane protrusion, cell migration and vesicle trafficking.

Here, we apply the structured-illumination microscopy (SIM) technique to analyse the actin cytoskeleton in fixed Drosophila Schneider (S2R+) cells, both in wild type and in cells depleted for WAVE, a major activator of Arp2/3 mediated actin polymerization. In addition, we demonstrate that live cell SIM imaging also allows the visualization of actin-driven lamellipodial membrane dynamics at high spatial resolution in S2R+ cells. Three dimensional (3D) SIM images of up to 70 μm thick Drosophila wild-type and abi-mutant egg chambers further enables us to resolve changes of actin structures in a multicellular context with an impressive lateral and axial resolution, which is not possible with conventional confocal microscopy. Thus, the combination of superresolution 3D microscopy with Drosophila genetics and cell biology allows detailed insights into the structural and molecular requirements of different actin-dependent processes.

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