Quantitative proteomics reveals insights into the assembly of IFT trains and ciliary assembly.

Intraflagellar transport (IFT) is required for ciliary assembly. The IFT machinery comprises IFT motors kinesin-2 and IFT dynein plus IFT-A and IFT-B complexes, which assemble into IFT trains in cilia. To gain mechanistic understanding of IFT and ciliary assembly, we have performed absolute quantification of IFT machinery in Chlamydomonas cilium. There are per cilium about 756, 532, 276, 350 molecules of IFT-B, IFT-A, IFT dynein and kinesin-2, respectively. The amount of IFT-B is sufficient to sustain rapid ciliary growth in terms of tubulin delivery. The stoichiometric ratio of IFT-B/IFT-A/dynein is about 3:2:1 whereas IFT-B/IFT-A ratio in an IFT dynein mutant is 2:1, suggesting a plastic interaction between IFT-A and IFT-B that can be influenced by IFT dynein. Considering diffusion of kinesin-2 during retrograde IFT, it is estimated that one kinesin-2 drives eight molecules of IFT-B during anterograde IFT. These data provide new insights into the assembly of IFT trains and ciliary assembly.