Cilia and flagella are highly conserved organelles that have diverse roles in cell motility and sensing extracellular signals. Motility defects in cilia and flagella often result in primary ciliary dyskinesia. However, the mechanisms underlying cilia formation and function, and in particular the cytoplasmic assembly of dyneins that power ciliary motility, are only poorly understood. Here we report a new gene, kintoun (ktu), involved in this cytoplasmic process. This gene was first identified in a medaka mutant, and found to be mutated in primary ciliary dyskinesia patients from two affected families as well as in the pf13 mutant of Chlamydomonas. In the absence of Ktu/PF13, both outer and inner dynein arms are missing or defective in the axoneme, leading to a loss of motility. Biochemical and immunohistochemical studies show that Ktu/PF13 is one of the long-sought proteins involved in pre-assembly of dynein arm complexes in the cytoplasm before intraflagellar transport loads them for the ciliary compartment.
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We thank C. Lo and D. Morris-Rosendahl for critical reading of this manuscript. We are grateful to M. Sugimoto, A. Ito-Igarashi, K. Nakaguchi, S. Minami, Y. H. Park, Y. Mochizuki, Y. Ozawa, K. Ohki, T. Obata, A. Heer and C. Reinhardt for excellent fish care and/or experimental assistance. We also thank A. Shimada and D. Nihei for their help in medaka experiments, J. Freshour and M. Nakatsugawa for help with Chlamydomonas, and S. King, H. Qin, W. Sale and D. Stern for antibodies. Our mutant screening was carried out mainly at the National Institute of Genetics (NIG), supported by NIG Cooperative Research Program (2002–2006). This work was supported in part by Grants-in-Aid for Scientific Research Priority Area Genome Science and Scientific Research (A and B), Global COE Program (Integrative Life Science Based on the Study of Biosignaling Mechanisms) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) of Japan, Yamada Science Foundation, and a Bio-Design Project of the Ministry of Agriculture, Forestry and Fisheries of Japan. D.K. was a research fellow supported by the 21th century COE program of the University of Tokyo, MEXT, Japan. This work was supported by grants to H.Omran from the ‘Deutsche Forschungsgemeinschaft’ DFG Om 6/4, GRK1104, BIOSS and the SFB592, and to D.R.M. from the NIH, GM44228. We would like to acknowledge the sequencing activities by K. Borzym and the Seq-Team at MPI-MG, which was supported by the German Ministry of Science and Education (BMBF) by grant NGFN-2:01GR0414-PDN-S02T17 to R.R. We are grateful for the support by the ‘Primare Ciliaere Dyskinesie and Kartagener Syndrom e.V.’.
Author Contributions Research planning and supervision was by H.Omran, D.R.M. and H.T.; medaka genetics and phenotypic analyses by D.K., T.T. and H.T.; biochemical experiments using mouse testis was by T.T., S.K. and Y.W.; high-speed video microscopy of medaka Kupffer’s vesicle cilia was by C.H., H.M., H.K., D.K. and A.M.; electron microscopy of medaka cilia/flagella was by H.H. and R.K.; experiments on human PCD were by H. Omran, H. Olbrich, N.T.L., M.F., H.Z., H.S. and R.R.; Chlamydomonas experiments were by D.R.M., Q.Z., G.L., E.O., T.Y. and R.K.; and manuscript writing was by H.Omran, D.R.M. and H.T.
Movie S1. Dorsal view of cilia in wild-type Kupffer's vesicle. The wild-type motile cilia rotate on the KV epithelial cells.
Movie S2. Dorsal view of cilia in ktu mutant Kupffer's vesicle. The cilia rotation is completely blocked.
Movie S3. Flagellar waveform of wild-type sperm. The wild-type flagellar bending beautifully propagate to the tip of the sperm tail.
Movie S4. Flagellar waveform of ktu mutant sperm. The mutant sperm looks paralyzed and the waveform of flagellar beating is affected. The flagellar bending does not propagate to the tip of the sperm tail.
Movie S5. Motility of cilia in respiratory cells from control patients.
Movie S6. Motility of cilia in respiratory cells from PCD patient OP146II1.
Movie S7. Motility of cilia in respiratory cells from PCD patient OP146II3.
Movie S8. Motility of cilia in respiratory cells from PCD patient OP234II1.
Movie S9. Motility of sperm flagella from control patients.
Movie S10. Motility of sperm flagella from PCD patient OP146II3.