In situ architecture of the ciliary base reveals the stepwise assembly of intraflagellar transport trains

The cilium is an antenna-like organelle that performs numerous cellular functions, including motility, sensing, and signaling. The base of the cilium contains a selective barrier that regulates the entry of large intraflagellar transport (IFT) trains, which carry cargo proteins required for ciliary assembly and maintenance. However, the native architecture of the ciliary base and the process of IFT train assembly remain unresolved. In this work, we used in situ cryo–electron tomography to reveal native structures of the transition zone region and assembling IFT trains at the ciliary base in Chlamydomonas . We combined this direct cellular visualization with ultrastructure expansion microscopy to describe the front-to-back stepwise assembly of IFT trains: IFT-B forms the backbone, onto which bind IFT-A, dynein-1b, and finally kinesin-2 before entry into the cilium. Cilia and flagella extend from the cell surface of various eukaryotic cells and perform diverse motility and signaling functions. The base of the cilium controls the entry of large intraflagellar transport trains that carry important cargo proteins throughout this specialized organelle. Defects in the ciliary base result in altered ciliary composition and human diseases. van den Hoek et al . used a combination of cryo–electron tomography and expansion microscopy techniques to study the molecular architecture of the ciliary base in the green alga Chlamydomonas reinhardtii . Their findings elucidate how intraflagellar transport trains assemble before they enter cilia and demonstrate the possibility of visualizing dynamic events with molecular resolution inside native cells. —SMH Native molecular structure of the Chlamydomonas ciliary transition zone and intraflagellar transport chain assembly is described.