ABSTRACT
Cilia are ubiquitous cellular organelles; the encoding ciliary genes are evolutionarily conserved from single-celled green algae to human. In addition to their roles as sensory antennae, cilia shed extracellular vesicles (EVs) that carry human ADPKD causal gene products polycystin-1 and polycystin-2 as cargos. The function of polycystin-carrying ciliary EVs is not known. The nematode C. elegans is a powerful animal model system for the study of PKD and human ciliopathies in the context of understanding the genetic control of ciliogenesis and ciliary function including EV biogenesis. Herein, we provide fundamental approaches and protocols used in the study of the polycystins and cilia using C. elegans as a model organism. Detailed protocols include PKD-2::GFP reporter EV quantification from single live worms, measuring in vivo cellular and intraflagellar transport (IFT) using fluorescently tagged reporters and time-lapse microscopy, RNAseq for cell-type specific transcriptome analysis, male mate search behavior analysis, behavioral assays as a readout for polycystin function in cilia and on EVs, preparing EV samples from C. elegans culture, and analysis EV morphology by negative staining and immunogold labeling. We aim to provide researchers—especially those new to the field—with sufficient background to initiate research using the C. elegans model to study human genetic diseases of cilia and EVs.
