Single depolymerizing and transport kinesins stabilize microtubule ends.

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.

Germanium nanospheres for ultraresolution picotensiometry of kinesin motors.

Kinesin motors are essential for the transport of cellular cargo along microtubules. How the motors step, detach, and cooperate with each other is still unclear. To dissect the molecular motion of kinesin-1, we developed germanium nanospheres as ultraresolution optical trapping probes.

The Kinesin-8 Kip3 Depolymerizes Microtubules with a Collective Force-Dependent Mechanism.

Microtubules are highly dynamic filaments with dramatic structural rearrangements and length changes during the cell cycle. An accurate control of the microtubule length is essential for many cellular processes, in particular during cell division. Motor proteins from the kinesin-8 family depolymerize microtubules by interacting with their ends in a collective and length-dependent manner.

Three-Dimensional Optical Tweezers Tracking Resolves Random Sideward Steps of the Kinesin-8 Kip3.

The budding yeast kinesin-8 Kip3 is a highly processive motor protein that walks to the ends of cytoskeletal microtubules and shortens them in a collective manner. However, how exactly Kip3 reaches the microtubule end is unclear. Although rotations of microtubules in multimotored Kip3 gliding assays implied directed sideward switching between microtubule protofilaments, two-dimensional, single-molecule, optical-tweezers assays indicated that Kip3 randomly switched protofilaments.

Phragmoplast Orienting Kinesin 2 Is a Weak Motor Switching between Processive and Diffusive Modes.

Plant development and morphology relies on the accurate insertion of new cell walls during cytokinesis. However, how a plant cell correctly orients a new wall is poorly understood. Two kinesin class-12 members, phragmoplast orienting kinesin 1 (POK1) and POK2, are involved in the process, but how these molecular machines work is not known.

Measuring Microtubule Supertwist and Defects by Three-Dimensional-Force-Clamp Tracking of Single Kinesin-1 Motors.

Three-dimensional (3D) nanometer tracking of single biomolecules provides important information about their biological function. However, existing microscopy approaches often have only limited spatial or temporal precision and do not allow the application of defined loads. Here, we developed and applied a high-precision 3D-optical-tweezers force clamp to track in vitro the 3D motion of single kinesin-1 motor proteins along microtubules.

Kinesin rotates unidirectionally and generates torque while walking on microtubules.

Cytoskeletal motors drive many essential cellular processes. For example, kinesin-1 transports cargo in a step-wise manner along microtubules. To resolve rotations during stepping, we used optical tweezers combined with an optical microprotractor and torsion balance using highly birefringent microspheres to directly and simultaneously measure the translocation, rotation, force, and torque generated by individual kinesin-1 motors.

Implementation and Tuning of an Optical Tweezers Force-Clamp Feedback System.

Feedback systems can be used to control the value of a system variable. In optical tweezers, active feedback is often implemented to either keep the position or tension applied to a single biomolecule constant. Here, we describe the implementation of the latter: an optical force-clamp setup that can be used to study the motion of processive molecular motors under a constant load.

The Kinesin-8 Kip3 switches protofilaments in a sideward random walk asymmetrically biased by force.

Molecular motors translocate along cytoskeletal filaments, as in the case of kinesin motors on microtubules. Although conventional kinesin-1 tracks a single microtubule protofilament, other kinesins, akin to dyneins, switch protofilaments. However, the molecular trajectory-whether protofilament switching occurs in a directed or stochastic manner-is unclear.