Coordination between Microtubule Motors during Bi-directional Cargo Transport in Live Cells.

摘要

Intracellular transport is typically bi-directional - consisting of a series of back and forth movements along a dense network of microtubules. Opposite-polarity molecular motors kinesin-1 and cytoplasmic dynein are interdependent in function during bi-directional transport of intracellular cargo i.e. inhibition or depletion of kinesin-1 abolishes dynein-driven cargo transport, and vice versa. However the mechanism mediating this interdependence remains unknown. In this work, we asked whether the basic mechanism of bi-directional intracellular transport requires any specific factor, other than the two oppositely-directed motors themselves i.e. is the presence of two oppositely-directed motors necessary and sufficient for bi-directional motility of organelles ? To this end, we determined whether any plus-end directed molecular motor can functionally replace kinesin-1 and conversely, whether any minus-end directed motor can functionally replace dynein in cargo transport.;Using Drosophila S2 cells, we demonstrated that replacement of endogenous kinesin-1 (kinesin heavy chain: KHC) or cytoplasmic dynein (dynein heavy chain: DHC) with an unrelated, peroxisome-targeted motor of the same directionality activated peroxisome transport in the opposite direction. Thus, replacement of endogenous Drosophila KHC with a fast and processive plus-end motor (dimerized C. elegans Unc104; Kinesin-3 family member) rescued bi-directional transport of peroxisomes. Even though slow and weakly processive motors activated the opposite-polarity motor, these motors could not compete with the endogenous motor. That is,;in an endogenous KHC-depleted background, expression of dimeric plus-end Xenopus Eg5 (Kinesin-5 family member) activated dynein-driven peroxisome transport resulting in accumulation of these organelles at the cell center, or minus-end. Similarly, in an endogenous DHC-depleted background, dimeric Drosophila Ncd (Kinesin-14 family member) activated KHC-driven peroxisome transport resulting in accumulation of these organelles at process tips, or the plus-ends. However motility-deficient versions of the above motors, which retain the ability to bind microtubules and hydrolyze ATP, did not activate peroxisome motility. Thus any pair of motors can activate one another provided they are (1) of the opposite-polarity (2) cargo-bound and (3) move along microtubules.;These results demonstrated that mechanical coordination between opposite-polarity motors is necessary and sufficient for bi-directional organelle transport. Finally, I present a 'mechanical signaling' model explaining these interactions between opposite-polarity motors during bi-directional cargo transport.