View Program - 63rd Annual Drosophila Research Conference

Over the course of animal development, distinct pools of epithelial cells are maintained by compartment boundaries. For example, in the early Drosophila embryo, the formation of compartment boundaries at regular intervals along the anterior-posterior axis establishes the segmental body plan of the future animal. These so called “parasegmental boundaries” (PSBs) are composed of highly stable cell-cell contacts that are under increased tension compared with typical contacts, and they resist the movement of cells between adjacent compartments. However, it is still unclear what differentiates PSB contacts from normal contacts at a molecular level. The force-generating cytoskeletal proteins F-actin and myosin are more highly enriched at PSBs compared with normal contacts, and evidence suggests that E-cadherin-based adherens junctions may also be differently arranged at PSBs. However, the fine details of these structures are below the resolution limit of standard confocal microscopy. To get past this resolution limit without the need for expensive super-resolution microscopes, we have adapted existing expansion microscopy (ExM) protocols for use in the early Drosophila embryo. ExM involves the isotropic physical expansion of a sample through chemical treatments to swell the embryo in a controlled manner, maintaining the relative positions of all molecules. We are using this technique to examine the ultrastructural distribution of adherens junction and cytoskeletal proteins at PSBs to see how they differ from normal contacts. We are also examining two known upstream regulators of PSB formation, Tartan and Ten-m, to better characterize their subcellular localization patterns in the early embryo. We expect these studies will give us a better understanding of how the three-dimensional arrangement of the cellular effectors of tension and adhesion––as well as the cell surface proteins that regulate them––gives PSBs their unique properties.

Using expansion microscopy to examine parasegmental boundaries at nanoscale resolution