Terminal cells lacking V-ATPase appear to form auto-cellular rather than seamless tubes. Deanne M. Francis, Amin Ghabrial. Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, PA.

   The tracheal system contains three tube architectures: multi-cellular tubes that have lumens surrounded by two or more cells; auto-cellular tubes that form when a single cell wraps around a lumenal space and seals itself into a tube by making self-junctions; and seamless tubes that form intra-cellularly generation of internal apical membrane surrounding a lumen. Tracheal terminal cells are found at the end of the branched tracheal network, where they ramify on muscle and internal tissues, making dozens of seamless tubes. Mutations in oak gall (okg) and conjoined (cnj) were identified in a forward genetic screen designed to uncover genes required for tracheal morphogenesis (Ghabrial,A.S. 2011). I have focused on the role of okg and cnj in tracheal terminal cells. Terminal cells mutant for okg or cnj share identical cell rounding, branch pruning, and local air-filling defects. Careful analysis has revealed that okg and cnj affect the distribution of E-Cadherin within the terminal cell, and in the most extreme cases alter the type of tube architecture present within the terminal cell such that tubes expected to be seamless instead appear to have auto-cellular junctions. Positional cloning revealed that okg encoded the E-subunit of the vacuolar (V)-ATPase and cnj encoded the G-subunit of the same multi-subunit complex. In fact, these two proteins hetero-dimerize (Ma et al., 2011). The V-ATPase holoenzyme acidifies intracellular organelles and is important for many cell processes (Nishi and Forgac. 2002). Interestingly, the V0 subunit of the V-ATPase has been shown to have an independent function in membrane fusion (Bayer et al, 2003). We find that depletion of V1 and V0 proteins cause similar phenotypes in tracheal terminal cells, suggesting that loss of the acidification function of the V-ATPase is responsible for the terminal cell phenotype. Current efforts towards distinguishing among possible models to explain the conversion of seamless to seamed tubes in okg and cnj mutant terminal cells will be presented.