Kinesin-binding–triggered conformation switching of microtubules contributes to polarized transport

Shima, Tomohiro; Morikawa, Manatsu; Kaneshiro, Junichi; Kambara, Taketoshi; Kamimura, Shinji; Yagi, Toshiki; Iwamoto, Hiroyuki; Uemura, Shunsuke; Shigematsu, Hideki; Shirouzu, Mikako; Ichimura, Taro; Watanabe, Tomonobu M.; Nitta, Ryo; Okada, Yasushi; Hirokawa, Nobutaka

doi:10.1083/jcb.201711178

Cited by 106 publications

(163 citation statements)

References 60 publications

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“…In detail, we find that for all 1 types of lattice that we tried, forward step dwell times are shorter, on average, than backstep 2 dwell times and detachment dwell times. This holds true also for median dwell times (not 3 shown). relative to a reference (see legend).…”

supporting

confidence: 52%

“…It is possible that rescued detachments might be a 1 more general principle of molecular motor function, important beyond the stepping 2 mechanism of kinesin-1. For example, for kinesins working in teams, rescued detachment of 3 the K.ADP-K.ADP state might serve to relieve internal strain within a team of collaborating 4 motors. 5 6 Mechanochemical coupling.…”

mentioning

confidence: 99%

“…MTs were spun down and resuspended in BRB80 supplemented with taxol (Sigma-1 Aldrich) or epothilone (VWR International) to 20 µM. In the case of unstabilised GDP MTs, 2 no drug was added but MTs were capped immediately following assembly using tubulin 3 premixed with 1 mM GMPCPP and incubated for 1 h, added to a final concentration of 1 4 µM. The capped MTs were spun down and resuspended in BRB80.…”

mentioning

confidence: 99%

“…(C) 5 Forward step (blue), backstep (red) and detachments (yellow) dwell times vs. hindering force 6 for Brain GDP MTs stabilised with taxol. The larger symbols are mean dwell times Probability of forward steps (blue), <12 nm backsteps (orange), >12 nm backsteps (maroon) 3 and detachments (yellow) versus force (pN) for kinesin stepping on different MT types. (E) 4 Relative probabilities for four types of event versus load, for stepping on 4 types of MT.…”

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confidence: 99%

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Backstepping mechanism of kinesin-1

Toleikis

Carter

Cross

2019

Preprint

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7Kinesins are molecular motors that walk along microtubules, hauling cargo through the 8 crowded cytoplasm of eukaryotic cells. At low load, kinesins almost always step towards 9 Significance statement 1 Kinesin-1 molecular motors typically walk towards microtubule plus ends. Under load 2 however, backsteps also occur. Backsteps have previously been thought to occur by 3 directional reversal of forwards walking. We report that kinesin waits before forward steps 4 for less time than before backsteps and detachments, that it waits for the same amount of time 5 before backsteps and detachments, and that the relative numbers of backsteps and 6 detachments can be shifted experimentally without affecting forward stepping. As we 7 discuss, this argues for a mechanism in which motors that fail to step forward within a time 8 window can slip back, re-engage, and retry forward stepping. Our findings speak to the 9 mechanisms of directional bias and mechanochemical coupling in kinesins, and potentially 10 other motors. 11 12

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supporting

confidence: 52%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Backstepping mechanism of kinesin-1

Toleikis

Carter

Cross

2019

Preprint

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“…However, mounting evidence suggests that kinesin binding to microtubules may result in conformational changes in the microtubule lattice that result in kinesin's cooperative binding [15][16][17] . Notably, GDP-microtubules elongate 1.3 -1.6 % when interacting with saturating concentrations of kinesin-1 18,19 . Implicit in these experiments and their interpretation is the concept of structural plasticity of the microtubule lattice playing a role in microtubule function 20 .…”

Section: Main Textmentioning

confidence: 99%

Modulation of kinesin’s load-bearing capacity by force geometry and the microtubule track

Pyrpassopoulos

Shuman

Ostap

2019

Preprint

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Kinesin motors and their associated microtubule tracks are essential for long-distance transport of cellular cargos 1 . Intracellular activity and proper recruitment of kinesins is regulated by biochemical signaling, cargo adaptors and microtubule associated proteins 2-4 . Here we report on a novel regulatory mechanism of kinesin's load-bearing capacity by forces across the microtubule track. Using optical tweezers and the three-bead assay 5,6 to specifically apply forces parallel to the long-axis of the microtubule, we found that the median attachment duration between kinesin and microtubules under opposing forces is up to 10-fold longer than observed previously using the more conventional single-bead assay, which is likely due to vertical forces imposed by the single bead 7 . Using the threebead assay, we also found that not all the protofilaments are equivalent interacting substrates for kinesin and that the median attachment duration of kinesin varies by more than 10-fold, depending on the relative angular position of the forces along the circumference of the microtubule. Thus, depending on the geometry of forces across the microtubule, kinesin can switch from a fast detaching motor (< 0.2 s) to a persistent motor that sustains attachment (> 3 s) at high forces (5 pN). Our data show that the load-bearing capacity of the kinesin motor is highly variable and can be dramatically affected by offaxis forces and forces across the microtubule lattice which has implications for a range of cellular activites including cell division and organelle transport.

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Single depolymerizing and transport kinesins stabilize microtubule ends

et al. 2021

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Microtubules are highly dynamic cellular filaments and an accurate control of their length is important for many intracellular processes like cell division. Among other factors, microtubule length is actively modulated by motors from the kinesin superfamily. For example, yeast kinesin-8, Kip3, motors depolymerize microtubules by a cooperative, force-and length-dependent mechanism. However, whether single motors can also depolymerize microtubules is unclear. Here, we measured how single kinesin motors influenced the stability of microtubules in an in vitro assay. Using label-free interference reflection microscopy, we determined the spontaneous microtubule depolymerization rate of stabilized microtubules in the presence of kinesins. Surprisingly, we found that both single Kip3 and nondepolymerizing kinesin-1 transport motors, used as a control, stabilized microtubules further. For Kip3, this behavior is contrary to the collective force-dependent depolymerization activity of multiple motors. Because of the control measurement, the finding may hint at a more general stabilization mechanism. The complex, concentration-dependent interaction with microtubule ends provides new insights into the molecular mechanism of kinesin-8 and its regulatory function of microtubule length.

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Kinesin-binding–triggered conformation switching of microtubules contributes to polarized transport

Cited by 106 publications

References 60 publications

Backstepping mechanism of kinesin-1

Backstepping mechanism of kinesin-1

Modulation of kinesin’s load-bearing capacity by force geometry and the microtubule track

Single depolymerizing and transport kinesins stabilize microtubule ends

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