Kashyap, S.; Xia, X.; Wang, H.; Shimogawa, M. M.; Agnew, A.; Hill, K.; Zhou, Z. H.

Axonemes are cylindrical bundles of microtubule filaments that typically follow a 9+n pattern (where n ranges from 0 to 4). However, variations exist across species and cell types, including architectures with fewer (e.g., 3+0, 6+0) or more than nine doublet microtubules (e.g., 9+9+0, 9+9+3), reflecting diverse structural adaptations of cilia and flagella in eukaryotes. Trypanosoma brucei, the causative agent of African trypanosomiasis, relies on its single 9+2 flagellum to navigate through environments within the mammalian host and insect vector. Central to the T. brucei flagellum\'s function is a canonical central apparatus (CA), composed of C1 and C2 singlet microtubules, which regulates flagellar beating and ensures efficient movement. Despite its crucial mechanoregulatory role in flagellar beating, the molecular structure and interactions governing T. brucei CA assembly and function remain poorly understood. In this study, we employed cryogenic electron microscopy (cryoEM) to uncover structural details of the T. brucei CA. We identified conserved and stably C1/C2-associated protein densities, including the armadillo repeat protein PF16, which serves as a structural scaffold critical for CA assembly and axonemal asymmetry. Our analysis also revealed pronounced molecular flexibility of the CA and uncovered T. brucei-specific densities, suggesting lineage-specific adaptations for parasite motility. These findings provide critical insights into the structural foundations of T. brucei motility. They also highlight potential therapeutic targets to disrupt the parasite\'s ability to cause disease, offering new avenues for the treatment of African trypanosomiasis. Comparison of CAs in this canonical 9+2 axoneme and non-canonical 9+n axonemes offers general insights into the assembly and diverse functions of CAs across a wide range of species.