Abstract
Intraflagellar transport (IFT), the key mechanism for ciliogenesis, involves large protein particles moving bi-directionally along the entire ciliary length. IFT particles contain two large protein complexes, A and B, which are constructed with proteins in a core and several peripheral proteins. Prior studies have shown that in Chlamydomonas reinhardtii, IFT46, IFT52, and IFT88 directly interact with each other and are in a subcomplex of the IFT B core. However, ift46, bld1, and ift88 mutants differ in phenotype as ift46 mutants are able to form short flagella, while the other two lack flagella completely. In this study, we investigated the functional differences of these individual IFT proteins contributing to complex B assembly, stability, and basal body localization. We found that complex B is completely disrupted in bld1 mutant, indicating an essential role of IFT52 for complex B core assembly. Ift46 mutant cells are capable of assembling a relatively intact complex B, but such complex is highly unstable and prone to degradation. In contrast, in ift88 mutant cells the complex B core still assembles and remains stable, but the peripheral proteins no longer attach to the B core. Moreover, in ift88 mutant cells, while complex A and the anterograde IFT motor FLA10 are localized normally to the transition fibers, complex B proteins instead are accumulated at the proximal ends of the basal bodies. In addition, in bld2 mutant, the IFT complex B proteins still localize to the proximal ends of defective centrioles which completely lack transition fibers. Taken together, these results revealed a step-wise assembly process for complex B, and showed that the complex first localizes to the proximal end of the centrioles and then translocates onto the transition fibers via an IFT88-dependent mechanism.
Highlights
Cilia and flagella are hair-like organelles extending from the basal bodies of almost all eukaryotic cells
We report that three complex B core proteins of Chlamydomonas have distinct roles in complex B assembly and stability and apical peri-basal body localization
By analyzing the intermediate complexes formed in mutants ift46, bld1, and ift88 (Fig. 1), this report clearly demonstrates that IFT46, IFT52, and IFT88, despite being in the same subcomplex [16], do not contribute to the assembly of complex B
Summary
Cilia and flagella are hair-like organelles extending from the basal bodies of almost all eukaryotic cells. These cellular appendages play important roles for cellular homeostasis and development. The assembly and function of the flagella relies on intraflagellar transport (IFT), the conserved bi-directional movement of linear trains of IFT particles along the axoneme [3]. Complex B is essential for the anterograde IFT, while complex A appears to mediate the retrograde movement [4,5,6,7,8]. In Chlamydomonas, both IFT A and B complexes contain a core complex and several peripheral proteins. The complex A core refers to the smaller complex found in a complex A mutant ift121 [11]. The complex B core is an 11S complex that stays intact in high ionic conditions [12]
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