Force generation of KIF1C is impaired by pathogenic mutations

Abstract

Intracellular transport is essential for neuronal function and survival. The most effective plus-end-directed neuronal transporter is the kinesin-3 KIF1C, which transports large secretory vesicles and endosomes.1Lipka J. Kapitein L.C. Jaworski J. Hoogenraad C.C. Microtubule-binding protein doublecortin-like kinase 1 (DCLK1) guides kinesin-3-mediated cargo transport to dendrites.EMBO J. 2016; 35: 302-318https://doi.org/10.15252/embj.201592929Crossref PubMed Scopus (86) Google Scholar, 2Schlager M.A. Kapitein L.C. Grigoriev I. Burzynski G.M. Wulf P.S. Keijzer N. de Graaff E. Fukuda M. Shepherd I.T. Akhmanova A. Hoogenraad C.C. Pericentrosomal targeting of Rab6 secretory vesicles by bicaudal-D-related protein 1 (BICDR-1) regulates neuritogenesis.EMBO J. 2010; 29: 1637-1651https://doi.org/10.1038/emboj.2010.51Crossref PubMed Scopus (109) Google Scholar, 3Lee P.L. Ohlson M.B. Pfeffer S.R. 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İşeri S.A.U. et al.Clinical phenotype of hereditary spastic paraplegia due to KIF1C gene mutations across life span.Brain Dev. 2018; 40: 458-464https://doi.org/10.1016/j.braindev.2018.02.013Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar, 7Caballero Oteyza A. Battaloğlu E. Ocek L. Lindig T. Reichbauer J. Rebelo A.P. Gonzalez M.A. Zorlu Y. Ozes B. Timmann D. et al.Motor protein mutations cause a new form of hereditary spastic paraplegia.Neurology. 2014; 82: 2007-2016https://doi.org/10.1212/WNL.0000000000000479Crossref PubMed Scopus (35) Google Scholar, 8Dor T. Cinnamon Y. Raymond L. Shaag A. Bouslam N. Bouhouche A. Gaussen M. Meyer V. Durr A. Brice A. et al.KIF1C mutations in two families with hereditary spastic paraparesis and cerebellar dysfunction.J. Med. Genet. 2014; 51: 137-142https://doi.org/10.1136/jmedgenet-2013-102012Crossref PubMed Scopus (50) Google Scholar In contrast to other kinesin-3s, KIF1C is a stable dimer and a highly processive motor in its native state.9Dorner C. Ullrich A. Häring H.-U. Lammers R. The kinesin-like motor protein KIF1C occurs in intact cells as a dimer and associates with proteins of the 14–3-3 family.J. Biol. Chem. 1999; 274: 33654-33660https://doi.org/10.1074/jbc.274.47.33654Abstract Full Text Full Text PDF PubMed Scopus (71) Google Scholar,10Siddiqui N. Zwetsloot A.J. Bachmann A. Roth D. Hussain H. Brandt J. Kaverina I. Straube A. PTPN21 and Hook3 relieve KIF1C autoinhibition and activate intracellular transport.Nat. Commun. 2019; 10: 2693https://doi.org/10.1038/s41467-019-10644-9Crossref PubMed Scopus (30) Google Scholar Here, we establish a baseline for the single-molecule mechanics of Kif1C. We show that full-length KIF1C molecules can processively step against the load of an optical trap and reach average stall forces of 3.7 pN. Compared with kinesin-1, KIF1C has a higher propensity to slip backward under load, which results in a lower maximal single-molecule force. However, KIF1C remains attached to the microtubule while slipping backward and re-engages quickly, consistent with its super processivity. Two pathogenic mutations, P176L and R169W, that cause hereditary spastic paraplegia in humans7Caballero Oteyza A. Battaloğlu E. Ocek L. Lindig T. Reichbauer J. Rebelo A.P. Gonzalez M.A. Zorlu Y. Ozes B. Timmann D. et al.Motor protein mutations cause a new form of hereditary spastic paraplegia.Neurology. 2014; 82: 2007-2016https://doi.org/10.1212/WNL.0000000000000479Crossref PubMed Scopus (35) Google Scholar,8Dor T. Cinnamon Y. Raymond L. Shaag A. Bouslam N. Bouhouche A. Gaussen M. Meyer V. Durr A. Brice A. et al.KIF1C mutations in two families with hereditary spastic paraparesis and cerebellar dysfunction.J. Med. Genet. 2014; 51: 137-142https://doi.org/10.1136/jmedgenet-2013-102012Crossref PubMed Scopus (50) Google Scholar maintain fast, processive single-molecule motility in vitro but with decreased run length and slightly increased unloaded velocity compared with the wild-type motor. Under load in an optical trap, force generation by these mutants is severely reduced. In cells, the same mutants are impaired in producing sufficient force to efficiently relocate organelles. Our results show how its mechanics supports KIF1C’s role as an intracellular transporter and explain how pathogenic mutations at the microtubule-binding interface of KIF1C impair the cellular function of these long-distance transporters and result in neuronal disease.

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