Abstract
Understanding the dynamic protein composition of cilia, crucial sensory organelles implicated in ciliopathies, is essential for comprehending cellular function and disease mechanisms. This Review summarizes advancements in methodologies for characterizing cilia proteomes. Traditional biochemical methods for isolating cilia from various model organisms have yielded informative protein inventories of cilia. These approaches worked particularly well for motile cilia but face challenges when investigating sparse primary cilia in various mammalian cells and tissues. The advent of protein-protein interaction mapping approaches employing genetically encoded affinity purification tags followed by mass spectrometry (AP-MS), has elucidated crucial ciliopathy-associated protein modules such as the BBSome, leading to new gene discoveries. More recently, proximity labeling techniques (APEX, BioID and TurboID) have revolutionized the field, enabling high-resolution and dynamic mapping of ciliary proteomes and signaling pathways, and dissecting protein trafficking defects in ciliopathy models. Despite technical variations, these methods offer novel insights into the ciliary and sub-ciliary protein environment. Further advancements in spatial proteomic technologies and mass spectrometry sensitivity might ultimately allow us to delineate dynamic proteomic profiles of specific cilia accurately across different cell types and tissues.