The structural heterogeneity of radial spokes in cilia and flagella is conserved

Abstract

Radial spokes are ubiquitous components of motile cilia and flagella and play an essential role in transmitting signals that regulate the activity of the dynein motors, and thus ciliary and flagellar motility. In some organisms the 96 nm axonemal repeat unit contains only a pair of two spokes, RS1 and RS2, while most organisms have spoke triplets with an additional spoke RS3. The spoke pair in Chlamydomonas flagella has been well characterized, while spoke triplets have received less attention. Here, we used cryo-electron tomography and subtomogram averaging to visualize the 3D structure of spoke triplets in Strongylocentrotus purpuratus (sea urchin) sperm flagella in unprecedented detail. Only small differences were observed between RS1 and RS2, but the structure of RS3 was surprisingly unique and structurally different from the other two spokes. We observed novel doublet specific features that connect RS2, RS3 and the nexin-dynein regulatory complex, three key ciliary and flagellar structures. The distribution of these doublet specific structures suggests that they could be important for establishing the asymmetry of dynein activity required for the oscillatory movement of cilia and flagella. Surprisingly, a comparison with other organisms demonstrated both that this considerable radial spoke heterogeneity is conserved and that organisms with radial spoke pairs contain the basal part of RS3. This conserved radial spoke heterogeneity may also reflect functional differences between the spokes and their involvement in regulating ciliary and flagellar motility.

Quantitative Proteomic Analysis of Human Airway Cilia Identifies Previously Uncharacterized Proteins of High Abundance

“…RTDR1, PPIL6, and NME5 were present at lower abundance. While several elegant studies (e.g., 14, 57, 58) have begun to determine the detailed molecular structure and composition of the radial spokes in other species, little is known about the relative abundance of individual radial spoke proteins in human cilia. Comparing the predicted stoichiometry of these proteins with those from a published model of a C. reinhardtii radial spoke (57) (Table 3) shows a similar pattern of abundance, with the spoke head proteins generally more abundant than those of the stalk or neck.…”

Computer‐assisted image analysis of human cilia and Chlamydomonas flagella reveals both similarities and differences in axoneme structure

“…Recent cryoET studies have demonstrated that while radial spoke 1 and 2 in Chlamydomonas are very similar, the third radial spoke in Tetrahymena and Strongylocentrotus is unique (Barber et al 2012; Lin et al 2012; Pigino et al 2011). The human average also shows that the third radial spoke is different in shape with a wider spoke head and a faint diagonal linker compared with spokes 1 and 2 (Figures 4B, D).…”

CMF22 Is a Broadly Conserved Axonemal Protein and Is Required for Propulsive Motility in Trypanosoma brucei

“…Of the 50 original CMF genes identified in T. brucei (19), 5 have now been demonstrated or implicated to function as part of the NDRC: trypanin, CMF70, CMF46, and CMF44 (22,23,25,26,64) and, as discussed above, CMF22. While CMF proteins themselves are broadly conserved, they interact with conserved as well as organism-specific proteins (94,95). Continued analysis of CMF22 and other CMF proteins therefore offers opportunities to enhance understanding of flagellum motility with relevance to the pathogenesis of trypanosomes and related human pathogens.…”