FIGURE 8:

RIB72BKO axonemes rescued with RIB72B-GFP show no structural defects and an additional tag density. (A–L) Tomographic slices (A–F) and isosurface renderings (G–L) in longitudinal views (in A, D, G, J proximal is on the right; in all remaining panels proximal is on the left) of Tetrahymena axonemal repeat units. The axonemal average from RIB72B-KO rescued with RIB72B-GFP (D-F, J-L) showed the same structures as seen in WT (A-C, G-I), that is, the mutant defects were rescued. Specifically, the loss of MIPs 1b, 4e, 6a, and 6d in the RIB72B-KO mutant (as seen in Figures 5H, empty green arrow; 6H, empty tomato arrow; and 7, G–I, empty teal and mauve arrows, respectively) is rescued by expression of RIB72B-GFP (green arrow and structure in D and J, tomato arrow and structure in L and F, and teal and mauve arrows and structures in E and K, respectively) and resemble the WT structures (green arrow in A, tomato arrow in C, and teal arrow in B, respectively). In addition, the extra density of the GFP-tag was visible in the vicinity of MIP4e in the rescued strain (green circles and densities in F and L) that was not observed in WT (compared with C and I). Slice positions of the tomographic slices are the same as in Figures 5 (for MIP1), 6 (for MIP4), and 7 (for MIP6); MIP structures are colored as follows: MIP1a blue, MIP1b green, MIP4a rose, MIP4b light orange, MIP4c dark orange, MIP4d yellow, MIP4e tomato, MIP6a teal, MIP6b aqua, MIP6c purple, MIP6d mauve. Other labels: A_t, A-tubule; B_t, B-tubule; I1, I1 dynein; IDA, inner dynein arms; MIP3; N-DRC, nexin dynein regulatory complex; ODA, outer dynein arms; RS, radial spokes; GFP, green fluorescent protein. Scale bar: 16 nm (valid A–F).