Cilia or eukaryotic flagella are microtubule-based organelles found across the eukaryotic tree of life. Their very high aspect ratio and crowded interior are unfavorable to diffusive transport of most components required for their assembly and maintenance. Instead, a system of intraflagellar transport (IFT) trains moves cargo rapidly up and down the cilium (Figure 1A).1-3 Anterograde IFT, from the cell body to the ciliary tip, is driven by kinesin-II motors, whereas retrograde IFT is powered by cytoplasmic dynein-1b motors.4 Both motors are associated with long chains of IFT protein complexes, known as IFT trains, and their cargoes.5-8 The conversion from anterograde to retrograde motility at the ciliary tip involves (1) the dissociation of kinesin motors from trains,9 (2) a fundamental restructuring of the train from the anterograde to the retrograde architecture,8,10,11 (3) the unloading and reloading of cargo,2 and (4) the activation of the dynein motors.8,12 A prominent hypothesis is that there is dedicated calcium-dependent protein-based machinery at the ciliary tip to mediate these processes.4,13 However, the mechanisms of IFT turnaround have remained elusive. In this study, we use mechanical and chemical methods to block IFT at intermediate positions along the cilia of the green algae Chlamydomonas reinhardtii, in normal and calcium-depleted conditions. We show that IFT turnaround, kinesin dissociation, and dynein-1b activation can consistently be induced at arbitrary distances from the ciliary tip, with no stationary tip machinery being required. Instead, we demonstrate that the anterograde-to-retrograde conversion is a calcium-independent intrinsic ability of IFT.

Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstraße 108 01307 Dresden Germany

Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstraße 108 01307 Dresden Germany; B CUBE Center for Molecular Bioengineering Technische Universität Dresden Tatzberg 41 01307 Dresden Germany; Cluster of Excellence Physics of Life Technische Universität Dresden 01062 Dresden Germany

Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstraße 108 01307 Dresden Germany; Center for Systems Biology Dresden Pfotenhauerstraße 108 01307 Dresden Germany

Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstraße 108 01307 Dresden Germany; Center for Systems Biology Dresden Pfotenhauerstraße 108 01307 Dresden Germany; Human Technopole 5 le Rita Levi Montalcini 1 20017 Milan Italy

Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstraße 108 01307 Dresden Germany; Cluster of Excellence Physics of Life Technische Universität Dresden 01062 Dresden Germany; Human Technopole 5 le Rita Levi Montalcini 1 20017 Milan Italy

Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstraße 108 01307 Dresden Germany; Institute of Molecular Genetics Czech Academy of Sciences Vídeňská 1083 142 20 Prague Czech Republic

UCSF School of Medicine 1550 4th Street San Francisco CA 94158 USA

Curr Biol. 2022 Sep 26;32(18):R967-R969. doi: 10.1016/j.cub.2022.07.061. PubMed

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