Tubular Excretory Canal Structure Depends on Intermediate Filaments EXC-2 and IFA-4 in Caenorhabditis elegans.

Abstract

The excretory canals of Caenorhabditis elegans are a model for understanding the maintenance of apical morphology in narrow single-celled tubes. Light and electron microscopy shows that mutants in exc-2 start to form canals normally, but these swell to develop large fluid-filled cysts that lack a complete terminal web at the apical surface, and accumulate filamentous material in the canal lumen. Here, whole-genome sequencing and gene rescue show that exc-2 encodes intermediate filament protein IFC-2 EXC-2/IFC-2 protein, fluorescently tagged via clustered regularly interspaced short palindromic repeats/Cas9, is located at the apical surface of the canals independently of other intermediate filament proteins. EXC-2 is also located in several other tissues, though the tagged isoforms are not seen in the larger intestinal tube. Tagged EXC-2 binds via pulldown to intermediate filament protein IFA-4, which is also shown to line the canal apical surface. Overexpression of either protein results in narrow but shortened canals. These results are consistent with a model whereby three intermediate filaments in the canals-EXC-2, IFA-4, and IFB-1-restrain swelling of narrow tubules in concert with actin filaments that guide the extension and direction of tubule outgrowth, while allowing the tube to bend as the animal moves.