No abstract

Kinesin-14 molecular motors represent an essential class of proteins that bind microtubules and walk toward their minus-ends. Previous studies have described important roles for Kinesin-14 motors at microtubule minus-ends, but their role in regulating plus-end dynamics remains controversial. Kinesin-14 motors have been shown to bind the EB family of microtubule plus-end binding proteins, suggesting that these minus-end–directed motors could interact with growing microtubule plus-ends. In this work, we explored the role of minus-end–directed Kinesin-14 motor forces in controlling plus-end microtubule dynamics. In cells, a Kinesin-14 mutant with reduced affinity to EB proteins led to increased microtubule lengths. Cell-free biophysical microscopy assays were performed using Kinesin-14 motors and an EB family marker of growing microtubule plus-ends, Mal3, which revealed that when Kinesin-14 motors bound to Mal3 at growing microtubule plus-ends, the motors subsequently walked toward the minus-end, and Mal3 was pulled away from the growing microtubule tip. Strikingly, these interactions resulted in an approximately twofold decrease in the expected postinteraction microtubule lifetime. Furthermore, generic minus-end–directed tension forces, generated by tethering growing plus-ends to the coverslip using λ-DNA, led to an approximately sevenfold decrease in the expected postinteraction microtubule growth length. In contrast, the inhibition of Kinesin-14 minus-end–directed motility led to extended tip interactions and to an increase in the expected postinteraction microtubule lifetime, indicating that plus-ends were stabilized by nonmotile Kinesin-14 motors. Together, we find that Kinesin-14 motors participate in a force balance at microtubule plus-ends to regulate microtubule lengths in cells.

Subjects with functional ankle instability present bilateral impairment of compensatory postural adjustments of the tibialis anterior in a support position and of the soleus of the uninjured limb regardless of the position. Subjects with mechanical instability present bilateral increase of these adjustments in the peroneal brevis regardless of the position and in the soleus muscle in the side of the perturbation.

Fluorescence (smTIRF) Microscopy. Contrary to the canonical theory, we determined the R5L mutation does not reduce Tau affinity for the microtubule. Rather, the R5L mutation reduces the total amount of Tau bound to the microtubule at saturating conditions. Our data suggests the R5L mutation reduces Tau:Tau interactions, decreasing the ability of R5L-Tau to form larger order complexes, known as ''Tau condensates.'' Currently, we are determining the mechanism by which this occurs, potentially due to a structural change associated with the R5L mutation. Altogether, these results challenge the current paradigm of how mutations in Tau lead to disease.