Fig. 3

Sub-flagellar localization of FAP174. (a) FAP174 Western blot of axonemes from WT and flagellar mutants. A polypeptide of ~10 kDa was present in the axonemes of WT and the radial spokes mutant pf14; whereas, among the central pair mutants it was reduced in the pf15 and pf20, and absent in pf18 and pf19 that lack the central pair microtubules. The presence of FAP174 in pf16 axoneme that lack the C1 microtubule suggest that FAP174 associates with the C2 microtubule. (b) The Ponceau-stained tubulin bands show equal loading of axonemal proteins. (c-e) Immunofluorescent microscopy of whole cells using pre-immune serum as a control to show autofluorescence and non-specificity of the antibody (images captured at 63x magnification). (f-q) Immunofluorescent microscopy of FAP174 comparing whole cells and nucleo-flagella apparatus (NFA) from wild type and pf18. The samples were decorated with anti-acetylated tubulin or anti-FAP174 as indicated. FAP174 localizes to the flagella as well as to the base of flagella possibly the transition zone (arrows). FAP174 was not detectable in the pf18 flagella but appeared as the bright spot at the base of flagella. (r-t) Localization of FAP174 around nucleus is more prominent in cells with low autofluorescence background and one such representative image is depicted here. The scale bars in the figure indicates 10 μm