Genetic dissection of tubulogenesis in the Drosophila trachea

Abstract: The formation of branched tubular organs, such as the mammalian lung kidney and vascular system, is an essential process in animal development. The Drosophila tracheal (respiratory) system provides a genetic model system to study the highly ordered process of branch outgrowth and fusion required to form continuous tubular networks.In a transposon mutagenesis screen we identified several genes involved in the different steps of tracheal network formation, including branch fusion. We initially studied the differentiation of the fusion cell in the dorsal branches (DBs). The DBs consists of 5-6 cells, one of which acquires the fusion cell fate. Our results point to a role for Decapentaplegic (Dpp), the Drosophila homolog of transforming growth factor-b (TGFb), in inducing the DB fusion cells fate. The Delta/Notch pathway is then required to select and restrict the fusion cell fate to a single cell in the DBs.After the outgrowth and fusion of tracheal branches, the constituent tubes acquire distinct size and shapes to generate a functional tubular tissue. We have found that a mutation in the grainyhead (grh) gene cause the branches to elongate excessively. Cellular and ultrastructural analyses showed that this phenotype was generated by apical cell membrane overgrowth. It was further shown that the activity of Grh is modulated by Branchless, the key regulator of branch outgrowth. Mutations in the Na/K ATPase a subunit (ATP a) and the fasiclin II (fasII) genes, cause similar elongated tracheal tubes as the ones found in grh mutants, but instead these mutations affect the lateral subcellular compartment, independently of the function of Grh in the apical cell domain.In a search for downstream effectors of Grh, we found the krotzkopf verkehrt (kkv) gene, encoding a Drosophila chitin synthase. Analysis of kkv mutants suggests that a chitinous intralumenal cable, lacking in kkv embryos, is essential for the tracheal tubes to attain their correct length and diameter. The composition of the chitinous filament is also affected in mutants of various septate junction components, indicating that their tube size defects are at least in part caused by an inability to correctly assemble the intralumenal chitinous matrix formed by Kkv.