Affiliations: 1) University of North Carolina at Chapel Hill, NC; 2) Syracuse University, NY
Keywords: r. cell-cell interactions; b. segmentation
Adherens junctions play a major role in assuring cell-cell adhesion, and through their linkage to the cytoskeleton, mediate dramatic shape changes and rearrangements during embryonic morphogenesis. The junctions need to be strong yet also dynamic, to allow cell rearrangement without tissue rupture. We want to understand how junction-associated proteins, like the multi-domain scaffold Polychaetoid (Pyd; fly homolog of ZO-1), work together to allow junctions to react dynamically to cell shape change and force generation. We hypothesize that the junctional complex and its linkage to the cytoskeleton involves multivalent connections among multiple junction-associated proteins, assembled into an underlying structured molecular complex that mediates binding to the cytoskeleton. I am using structured illumination microscopy (SIM) to explore the junctional localization of these proteins in high resolution. My preliminary data suggest that the core junction-associated proteins Canoe (Cno) and Armadillo (β-Catenin) localize to alternating clusters along the zonula adherens of cell-cell borders, often with little overlap. Furthermore, I found that Pyd surrounds these core junctional proteins in a cloud-like pattern of puncta. I am now broadening my analysis to include other junction-associated proteins like Bazooka (Par-3), and Non-muscle myosin II. Next, I will analyze how these patterns change in pyd mutants. In parallel, I am exploring the functional role of Pyd as one player in this robust network. While Pyd is not absolutely essential for embryonic viability, mutants display defects in cell rearrangements during germband extension. Mutants have stacks of elongated cells, an elevated number of rosettes and junctional disruptions at tricellular junctions and anterior-posterior cell borders, suggesting defects in responding to elevated force on cell-cell junctions. However, these occur without major tissue rupture, suggesting other proteins act in parallel. To explore a hypothesized change in junction dynamics in pyd embryos, I am applying live imaging during germband elongation. Furthermore, we are now expanding our analysis to include other players in this protein network, including the tricellular junction protein Sidekick, and Canoe’s regulators Rap1 and the GEF Dizzy.