Intrinsically disordered domain of kinesin-3 Kif14 enables unique functional diversity

Abstract

In addition to their force-generating motor domains, kinesin motor proteins feature various accessory domains enabling them to fulfil a variety of functions in the cell. Human kinesin-3, Kif14, localizes to the midbody of the mitotic spindle and is involved in the progression of cytokinesis. The specific motor properties enabling Kif14's cellular functions, however, remain unknown. Here, we show in vitro that it is the intrinsically disordered N-terminal domain of Kif14 that enables unique functional diversity of the motor. Using single molecule TIRF microscopy we observed that the presence of the disordered domain i) increased the Kif14 run-length by an order of magnitude, rendering the motor super-processive and enabling the motor to pass through highly crowded microtubule areas shielded by cohesive layers of microtubule-associated protein tau, which blocks less processive motors ii) enabled robust, autonomous Kif14 tracking of growing microtubule tips, independent of microtubule end-binding (EB) proteins and iii) enabled Kif14 to crosslink parallel microtubules and to drive the relative sliding of antiparallel ones. We explain these features of Kif14 by the observed increased affinity of the disordered domain for GTP-like tubulin and the observed diffusible interaction of the disordered domain with the microtubule lattice. We hypothesize that the disordered domain tethers the motor domain to the microtubule forming a diffusible foothold. We suggest that the intrinsically disordered N-terminal anchoring domain of Kif14 is a regulatory hub supporting the various cellular functions of Kif14 by tuning the motor's interaction with microtubules.

VASH1-SVBP and VASH2-SVBP generate different detyrosination profiles on microtubules

“…In addition, VASH1 might have a higher cleavage efficiency than VASH2, as suggested by the increased detyrosination activity of the central domain of VASH1 compared to the one of VASH2 (Figure 5B). Notably, our work reveals that as for molecular motors (Zhernov et al, 2020) electrostatic interactions are key factors in the association between VASH-SVBP complexes and microtubules, making their processive enzymatic functioning possible. Indeed, the disordered flanking regions of the highly structured VASH core domains fine-tune the association via such electrostatic interactions.…”

“…Notably, our work reveals that as for molecular motors (Zhernov et al, 2020) electrostatic interactions are key factors in the association between VASH-SVBP complexes and microtubules, making their processive enzymatic functioning possible. Indeed, the disordered flanking regions of the highly structured VASH core domains fine-tune the association via such electrostatic interactions.…”

Reconstitution of an active human CENP-E motor

“…The increase in processivity may also be due to the presence of a non-motor microtubule-binding site at the far C terminus of full-length CENP-E, as CENP-E 754 -2mNeon lacks this region [28][29][30]. Many kinesins have a second non-specific microtubule-binding site, which increases their residency time and processivity [31][32][33].…”

Fast leaps between millisecond confinements govern Ase1 diffusion along microtubules

“…Diffusion is one of the essential strategies that proteins use to navigate the complex environment of the MT surface. It is employed by a large variety of MAPs, including molecular motors, 1,[40][41][42] intrinsically disordered neuronal proteins, 9,[43][44][45] proteins tracking polymerizing or depolymerizing microtubule tips 4,9,46,47 or microtubule crosslinker proteins. 4,9,15,48 Stepping of proteins from the Ase1/PRC1/MAP65 family across protofilaments has been predicted theoretically 9 , and here we present a direct experimental validation of this prediction.…”