Rowsell, C. M.; Kubo, S.; Arin, A.; Legal, T.; Yu, Y.; Bui, K. H.

Centrioles are essential cellular organelles with complex architectures. Their earliest assembly intermediate, the cartwheel, has a 9-fold symmetry established by Spindle assembly abnormal protein 6 (SAS-6). We used cryo-electron tomography and sub-tomogram averaging to resolve the native structure of the SAS-6 rings within the cartwheel of the exceptionally long Trichonympha proximal centriole. A 16-nm axial periodicity was formed by stacked rings of V-shaped SAS-6 tetramers, which formed the fundamental unit of the cartwheel's central hub. SAS-6 head domains adopted a zigzag stacking pattern. Furthermore, the improved resolution provides new insights into the function of the previously observed central inner domain, which forms nine asymmetric densities that bridge adjacent tetramers, thereby imparting polarity and enhancing structural stability. Molecular dynamics simulations indicate increased rigidity in inter-tetramer interactions, supporting efficient ring formation and revealing how cartwheel architecture and polarity are established and stabilized.