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IFT54 regulates IFT20 stability but is not essential for tubulin transport during ciliogenesis

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Abstract

Intraflagellar transport (IFT) is required for ciliogenesis by ferrying ciliary components using IFT complexes as cargo adaptors. IFT54 is a component of the IFT-B complex and is also associated with cytoplasmic microtubules (MTs). Loss of IFT54 impairs cilia assembly as well as cytoplasmic MT dynamics. The N-terminal calponin homology (CH) domain of IFT54 interacts with tubulins/MTs and has been proposed to transport tubulin during ciliogenesis, whereas the C-terminal coiled-coil (CC) domain binds IFT20. However, the precise function of these domains in vivo is not well understood. We showed that in Chlamydomonas, loss of IFT54 completely blocks ciliogenesis but does not affect spindle formation and proper cell cycle progression, even though IFT54 interacts with mitotic MTs. Interestingly, IFT54 lacking the CH domain allows proper flagellar assembly. The CH domain is required for the association of IFT54 with the axoneme but not with mitotic MTs, and also regulates the flagellar import of IFT54 but not IFT81 and IFT46. The C-terminal CC domain is essential for IFT54 to bind IFT20, and for its recruitment to the basal body and incorporation into IFT complexes. Complete loss of IFT54 or the CC domain destabilizes IFT20. ift54 mutant cells expressing the CC domain alone rescue the stability of IFT20 and form stunted flagella with accumulation of both IFT-A component IFT43 and IFT-B component IFT46, indicating that IFT54 also functions in IFT turn-around at the flagellar tip.

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Abbreviations

CH domain:

Calponin homology domain

CrIFT54:

Chlamydomonas IFT54

CC domain:

Coiled-coil domain

IFT:

Intraflagellar transport

DIC:

Differential interference contrast microscopy

Microtubules:

MTs

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Acknowledgements

We thank Dr. Roger Sloboda for critical review of this manuscript. We are also grateful to Dr. Karl Lechtreck for sharing unpublished data, and Drs. Joel Rosenbaum, Dennis Diener, George Witman, Robert Bloodgood and Kaiyao Huang for providing antibodies and plasmids. This work was supported by the following awards (to J.P.) from the National Basic Research Program of China (973 program) (2013CB910700), National Natural Science Foundation of China (31330044, 31671387) and Sino-German Center for Research Promotion (GZ990).

Author contributions

XZ., Y.L. and F.G. designed and performed the experiments and analyzed the data. J.P. designed the study, analyzed the data and wrote the paper together with X.Z.

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Authors and Affiliations

  1. MOE Key Laboratory of Protein Sciences, Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China

    Xin Zhu, Yinwen Liang, Feng Gao & Junmin Pan

  2. Developmental Biology Program, Sloan Kettering Institute, New York, NY, 10065, USA

    Yinwen Liang

  3. Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, Shandong Province, China

    Junmin Pan

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  1. Xin Zhu
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Correspondence to Junmin Pan.

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Zhu, X., Liang, Y., Gao, F. et al. IFT54 regulates IFT20 stability but is not essential for tubulin transport during ciliogenesis. Cell. Mol. Life Sci. 74, 3425–3437 (2017). https://doi.org/10.1007/s00018-017-2525-x

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