Mutants with reduced expression of FAP74 lack the C1d central pair projection. (A) Diagram of FAP74 artificial miRNA construct. (B) Western blot of WT and mutant flagella showing that FAP74 amiRNA transformants 1G11, 2D4, and 7A4 have reduced levels of FAP74. FAP221 levels are not reduced in these mutants. IC138 is a dynein intermediate chain used as a loading control. (C) Top panel is a silver-stained gel of high calcium anti-CaM IPs from axonemal extracts. Reduced amounts of FAP54 and FAP46 are immunoprecipitated from 1G11, 2D4, and 7A4 axonemal extracts compared with WT. Bottom panels are anti-FAP74 and anti-FAP221 Western blots of high calcium anti-CaM IPs showing that FAP74 and FAP221 are not precipitated from FAP74ami transformants. (D) Diagram shows the identities of the central pair projections. Electron micrographs of transverse sections of WT, 1G11, pf14, and 1G11,pf14 mutant axonemes. (E) Enlarged view of the central apparatus from pf14 and 1G11,pf14 double-mutant axonemes. All micrographs are oriented with the axoneme viewed proximal to distal with the bar representing 25 nm; the C1 central microtubule in all images is to the left. The 1G11 and 1G11,pf14 axonemes lack the C1d density and the sheath connecting C1d to C1b (marked by red arrows and asterisks).