A global analysis of IFT-A function reveals specialization for transport of membrane-associated proteins into cilia

Abstract

Intraflagellar transport (IFT), trafficking within the cilium of trains of IFT particles carrying cargos, is essential for formation of cilia in most organisms. Defects in IFT cause several human diseases. IFT trains are composed of two complexes, IFT-A and IFT-B. To dissect the functions of these complexes, we studied a Chlamydomonas mutant null for the IFT-A protein IFT140. The mutation had no effect on IFT-B but destabilized IFT-A, preventing flagella assembly. Therefore, IFT-A assembly requires IFT140. Truncated IFT140, lacking the protein's N-terminal WD repeats, partially rescued IFT and supported formation of half-length flagella containing normal levels of IFT-B but greatly reduced amounts of IFT-A. The axonemes of these flagella had normal ultrastructure, and by SDS-PAGE the composition of the axoneme appeared normal but that of the flagellar “membrane+matrix” was abnormal. Analysis of the latter fraction by mass spectrometry revealed decreases in small GTPases, lipid-anchored proteins, and cell signaling proteins. Thus, IFT-A is specialized for the import of membrane-associated proteins. Abnormal levels of the latter likely account for the multiple phenotypes of patients with defects in IFT140.