TY - JOUR
T1 - Kinesin-2 from C. reinhardtii Is an Atypically Fast and Auto-inhibited Motor that Is Activated by Heterotrimerization for Intraflagellar Transport
AU - Sonar, Punam
AU - Youyen, Wiphu
AU - Cleetus, Augustine
AU - Wisanpitayakorn, Pattipong
AU - Mousavi, Sayed I.
AU - Stepp, Willi L.
AU - Hancock, William O.
AU - Tüzel, Erkan
AU - Ökten, Zeynep
N1 - Funding Information:
This work was supported by the European Research Council grant 335623 to Z.Ö. and the National Institute of Health grants nos. R01GM100076 and R01GM121679 to W.O.H. and E.T. We acknowledge the Thai government for supporting W.Y. and P.W. through the Development and Promotion of Science and Technology (DPST) Scholarship. The authors would also like to thank all members of the Ökten and Tüzel groups for helpful discussions and their insightful suggestions. P.S. and Z.Ö. designed the experiments. P.S. and A.C. performed experiments and analyzed the data. W.L.S. wrote all customized MATLAB routines. W.Y. developed the model, performed the simulations, analyzed data, and prepared the figures; P.W. analyzed data and prepared figures; S.I.M. developed the Monte Carlo simulations for calculating the landing distributions; and E.T. supervised all the modeling work. Z.Ö. P.S. W.O.H. W.Y. P.W. and E.T. contributed to the manuscript writing. The authors declare no competing interests.
Funding Information:
This work was supported by the European Research Council grant 335623 to Z.Ö. and the National Institute of Health grants nos. R01GM100076 and R01GM121679 to W.O.H. and E.T. We acknowledge the Thai government for supporting W.Y. and P.W. through the Development and Promotion of Science and Technology (DPST) Scholarship. The authors would also like to thank all members of the Ökten and Tüzel groups for helpful discussions and their insightful suggestions.
Publisher Copyright:
© 2020 Elsevier Ltd
PY - 2020/3/23
Y1 - 2020/3/23
N2 - Construction and function of virtually all cilia require the universally conserved process of intraflagellar transport (IFT) [1, 2]. During the atypically fast IFT in the green alga C. reinhardtii, on average, 10 kinesin-2 motors “line up” in a tight assembly on the trains [3], provoking the question of how these motors coordinate their action to ensure smooth and fast transport along the flagellum without standing in each other's way. Here, we show that the heterodimeric FLA8/10 kinesin-2 alone is responsible for the atypically fast IFT in C. reinhardtii. Notably, in single-molecule studies, FLA8/10 moved at speeds matching those of in vivo IFT [4] but additionally displayed a slow velocity distribution, indicative of auto-inhibition. Addition of the KAP subunit to generate the heterotrimeric FLA8/10/KAP relieved this inhibition, thus providing a mechanistic rationale for heterotrimerization with the KAP subunit fully activating FLA8/10 for IFT in vivo. Finally, we linked fast FLA8/10 and slow KLP11/20 kinesin-2 from C. reinhardtii and C. elegans through a DNA tether to understand the molecular underpinnings of motor coordination during IFT in vivo. For motor pairs from both species, the co-transport velocities very nearly matched the single-molecule velocities, and both complexes spent roughly 80% of the time with only one of the two motors attached to the microtubule. Thus, irrespective of phylogeny and kinetic properties, kinesin-2 motors work mostly alone without sacrificing efficiency. Our findings thus offer a simple mechanism for how efficient IFT is achieved across diverse organisms despite being carried out by motors with different properties.
AB - Construction and function of virtually all cilia require the universally conserved process of intraflagellar transport (IFT) [1, 2]. During the atypically fast IFT in the green alga C. reinhardtii, on average, 10 kinesin-2 motors “line up” in a tight assembly on the trains [3], provoking the question of how these motors coordinate their action to ensure smooth and fast transport along the flagellum without standing in each other's way. Here, we show that the heterodimeric FLA8/10 kinesin-2 alone is responsible for the atypically fast IFT in C. reinhardtii. Notably, in single-molecule studies, FLA8/10 moved at speeds matching those of in vivo IFT [4] but additionally displayed a slow velocity distribution, indicative of auto-inhibition. Addition of the KAP subunit to generate the heterotrimeric FLA8/10/KAP relieved this inhibition, thus providing a mechanistic rationale for heterotrimerization with the KAP subunit fully activating FLA8/10 for IFT in vivo. Finally, we linked fast FLA8/10 and slow KLP11/20 kinesin-2 from C. reinhardtii and C. elegans through a DNA tether to understand the molecular underpinnings of motor coordination during IFT in vivo. For motor pairs from both species, the co-transport velocities very nearly matched the single-molecule velocities, and both complexes spent roughly 80% of the time with only one of the two motors attached to the microtubule. Thus, irrespective of phylogeny and kinetic properties, kinesin-2 motors work mostly alone without sacrificing efficiency. Our findings thus offer a simple mechanism for how efficient IFT is achieved across diverse organisms despite being carried out by motors with different properties.
UR - http://www.scopus.com/inward/record.url?scp=85081661819&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85081661819&partnerID=8YFLogxK
U2 - 10.1016/j.cub.2020.01.046
DO - 10.1016/j.cub.2020.01.046
M3 - Article
C2 - 32142698
AN - SCOPUS:85081661819
SN - 0960-9822
VL - 30
SP - 1160-1166.e5
JO - Current Biology
JF - Current Biology
IS - 6
ER -