Macromolecular crowding regulates the velocity of intracellular transport by kinesin-1 motors*

The cytosol is crowded with a high concentration of macromolecules. Crowding can alter protein conformation, binding rates, and reaction kinetics, yet it is not known how crowding affects intracellular cargo transport by molecular motor proteins. We use live cell and single-molecule imaging and optical trap measurements to uncover the consequences of macromolecular crowding on cargo transport by kinesin-1 motors. Surprisingly, we find that crowding significantly slows transport by teams of motors, while having no effect on single motor velocity. We find that this emergent property of kinesin teams results from the individuals’ increased sensitivity to hindering load in a crowded medium. Our results highlight the importance of motor-motor interactions in cargo transport, explain the long observed variability of cargo velocity, and suggest the use of crowding as a control parameter to study kinesin’s mechanochemical cycle.