Figure 5.

Analysis of TBB-4::YFP transport rate in cilia. Kymographs of DYF-1::GFP and TBB-4::YFP in IFT assays under exactly same conditions except that TBB-4::YFP was photobleached with a mercury lamp before recording to reduce the background. (a) DYF-1::GFP represents the IFT transport in cilia, and the IFT tracks are clear and thick in the kymograph. (b) The tracks of TBB-4::YFP in cilia are faint and thin compared to IFT tracks, e.g. in (a). (c) For comparison, OSM-9::GFP, which is proposed to be transported by IFT was used as control. All the recorded movies were processed using the basic filters (Sharpen (High) and Low pass) before creating kymographs. K is the kymograph that was created along the cilia and K′ is a drawing of the kymograph lines in K. In (a–c), Horizontal bar=2.5 μm and vertical bar=5 s. (d–f) Modeling of MT dynamics in a cilium. Dynamic instability and the delivery of tubulin subunits via IFT can constrain the length fluctuations of MTs in both the middle (blue) and the distal (black) segments to a narrow range (d). In silico FRAP of the cilium shown in (d) for both the middle (e) and distal (f) segments indicates similar recovery curves to the experimental results (Figure 4d, e). The fluorescence intensity is normalized to the prebleach. The labels are: DS, distal segment; MS, middle segment.