The excretory canal cell is the largest single epithelial cell within C. elegans, and its long fluid-filled hollow canals forms a useful model for understanding single-cell tubulogenesis. Our previous studies have identified a set of exc genes whose products regulate and maintain the narrow diameter of the canals during outgrowth. Some of these genes encode cytoskeletal proteins (including beta Hspectrin, formin, and intermediate filament proteins), and vesicular transport proteins (CRIP, IRG, and FGD proteins). The Miller lab has made available a list of genes highly enriched in various tissues of C. elegans (www.vanderbilt.edu/wormdoc/wormmap/WormViz.html), including genes enriched in the excretory canal cell. We have gratefully built on these results to search for additional genes that affect excretory canal structure, as well as genes that interact with known exc genes. We designed and are conducting a feeding RNAi screen with ~200 candidate genes strongly expressed in the canal cell, and which encode proteins with good potential to interact with EXC proteins, with the purpose of understanding the network of structural and trafficking required to build and maintain a narrow tube in a growing, moving animal. Since the excretory canal cell may be refractory to RNAi, as it has some neural characteristics (long processes following neural guidance cues during outgrowth), we crossed our chosen exc strains to an RNAi-sensitive strain (
rrf-3) that carries an integrated construct marking the canal cell cytoplasm (Pvha-1::gfp). The selected exc mutant backgrounds represent several pathways of interest: cytoskeletal structure (
exc-2), vesicular transport (
exc-5 and
exc-9) , RNA-binding (
exc-7), and ion transport (
exc-4), as well as animals with wild-type canals. At present, we have isolated and verified close to ten hits, some of which exacerbate canal defects, and excitingly, one which at least partially rescues the strongly cystic phenotype of the
exc-5 mutant. These knockdowns offer insight into novel gene interactions as well as alternative pathways that have yet to be explored in regards to excretory canal maintenance. We are continuing the RNAi screen.