The Triple-Repeat Protein Anakonda Controls Epithelial Tricellular Junction Formation in Drosophila

• Bicellular and tricellular occluding junctions form independently in Drosophila
• The transmembrane protein Anakonda localizes to epithelial tricellular junctions
• Anakonda is crucial for epithelial barrier function and tricellular junction assembly
• Anakonda promotes the accumulation of Gliotactin at tricellular junctions

SummaryIn epithelia, specialized tricellular junctions (TCJs) mediate cell contacts at three-cell vertices. TCJs are fundamental to epithelial biology and disease, but only a few TCJ components are known, and how they assemble at tricellular vertices is not understood. Here we describe a transmembrane protein, Anakonda (Aka), which localizes to TCJs and is essential for the formation of tricellular, but not bicellular, junctions in Drosophila. Loss of Aka causes epithelial barrier defects associated with irregular TCJ structure and geometry, suggesting that Aka organizes cell corners. Aka is necessary and sufficient for accumulation of Gliotactin at TCJs, suggesting that Aka initiates TCJ assembly by recruiting other proteins to tricellular vertices. Aka’s extracellular domain has an unusual tripartite repeat structure that may mediate self-assembly, directed by the geometry of tricellular vertices. Conversely, Aka’s cytoplasmic tail is dispensable for TCJ localization. Thus, extracellular interactions, rather than TCJ-directed intracellular transport, appear to mediate TCJ assembly.