Cryo-electron tomography of eel sperm flagella reveals a molecular "minimum system" for motile cilia

Cilia and flagella play a crucial role in the development and function of eukaryotes. The activity of thousands of dyneins is precisely regulated to generate flagellar motility. The complex proteome (600+ proteins) and architecture of the structural core of flagella, the axoneme, have made it challenging to dissect the functions of the different complexes, like the regulatory machinery. Previous reports suggested that the flagellum of American eel sperm lacks many of the canonical axonemal complexes yet is still motile. Here, we use cryo-electron tomography for molecular characterization of this proposed "minimal" motile flagellum. We observed different diameters for the eel sperm flagellum: narrow at the base and wider toward the flagellar tip. Subtomogram averaging revealed the three-dimensional (3D) structure of the eel sperm flagellum. As expected, major complexes were missing, for example, outer dynein arms, radial spokes, and the central pair complex, but we found molecular remnants of most complexes. We also identified bend direction-specific patterns in the inter-DMT distance in actively beating eel sperm flagella and we propose a model for the regulation of dynein activity during their motility. Together, our results shed light on the structure and function of the eel sperm flagellum and provide insight into the minimum requirements for ciliary beating. SIGNIFICANCE STATEMENT center dot Regulating dyneins is important for flagellar beating. The [9+0] eel sperm flagellum lacks many structures responsible for dynein regulation and the molecular mechanism of eel sperm motion is unknown. center dot The authors characterized the 3D structures of the eel sperm flagella using cryo-electron tomography and found molecular remnants of several axonemal complexes that were thought to be missing, including radial spokes and outer dynein arms. They also identified unique features, like variable inter-DMT distances, which may play roles in generating flagellar beating. center dot These findings reveal a minimal system for motile flagella, providing insights into the basic requirements and functions of ciliary complexes.