UC Berkeley

Kinesin and dynein are motor proteins that carry intracellular cargos on microtubule filaments (MTs). The regulation of these motors is critical for the internal organization of cells, as defects in transport cause aberrant mitosis, neurodevelopmental and neurodegenerative disease, and cancer. We have a strong mechanistic understanding of how motors utilize the energy of ATP hydrolysis to walk unidirectionally along MTs, but it remains unclear how they are regulated in the complex intracellular environment.MT associated proteins (MAPs) bind and organize the MT network and regulate the motors that transport cellular cargos along these tracks. While most MAPs play an inhibitory role on motors, MAP7 enhances kinesin-1-driven transport, but the mechanism remains unclear. We mechanistically dissected the interactions of MAP7 and kinesin-1 using a combination of biochemistry, structural biology, and fluorescence microscopy. Through cryo-EM, our collaborators found that the MAP7 and kinesin-1 MT binding sites directly overlap, and we showed that this overlap causes inhibition of motor binding and activity. The MAP7 projection domain recruited kinesin-1 to the MT and increases its motile parameters. These two opposing functions of MAP7 lead to a biphasic, concentration-dependent regulation of kinesin-1, where the activating component dominates at low MAP7 concentrations, but the inhibiting component is dominantat high MT coverage.I next studied how MAPs and motors are regulated by MT tracks. MTs are heavily post-translationally modified at the carboxy-terminal tails of tubulin subunits, and the MT lattice switches between extended and compacted conformations. It remained unclear how tubulin modifications or lattice conformation play a role in MAP-motor interactions. We found that kinesin-1 is mostly insensitive to tubulin modifications, but its activity is tuned by MT lattice spacing. Similarly, a subset of MAPs (tau, MAP7 and doublecortin) is highly sensitive to MT lattice spacing but mostly insensitive to tubulin PTMs. We showed how tau reads and writes the MT lattice spacing, and how this impacts the competition between MAP7 and tau to decorate MTs. In turn, MT decoration by these MAPs dictates whether kinesin-1 can walk along those tracks. These results suggest that cells use a two-layered regulatory code: lattice spacing of MTs changes the MAP decoration of MTs, and MAPs regulate kinesin-1 activity to control intracellular traffic.