Measuring rates of intraflagellar transport along Caenorhabditis elegans sensory cilia using fluorescence microscopy

Intraflagellar transport (IFT), the kinesin-2 and IFT-dynein-dependent bidirectional movement of multisubunit protein complexes called IFT-particles and associated cargo molecules along ciliary axonemes, is thought to be essential for the assembly and maintenance of virtually all eukaryotic cilia and flagella. Transport assays that allow measurements of the rates of movement of specific, fluorescently tagged, functional components of the IFT machinery, including motors, IFT particle subunits, and putative cargo, were first developed in Caenorhabditis elegans sensory cilia, and they have proved to be an important and valuable tool for dissecting the molecular mechanisms of IFT. We describe how these transport assays are performed in our laboratory and summarize the information that has been obtained by using them concerning the mechanisms of action and regulation of the motors that drive IFT, the composition and organization of the IFT-particles, and the identification of IFT-dynein subunits and ciliary tubulin isotypes as likely cargo proteins of kinesin-2-driven anterograde IFT.