Kinesin Regulation: Discovering the Mechanisms that Mediate Autoinhibition, Cargo Complex Formation, and Selective Microtubule Use in Neurons.

Abstract

Kinesin motors play an indispensable role in intracellular trafficking by driving the long-distance transport of protein and vesicular cargoes along microtubules. My thesis work has focused on uncovering regulatory mechanisms that mediate 1) kinesin autoinhibition, 2) cargo complex formation, and 3) polarized transport due to preferential microtubule track selection. Kinesin motors generally are kept inactive in the absence of cargo. My work demonstrates that KIF17 (Kinesin-2) is self-inhibited by assuming a folded conformation that allows a weak coiled-coil region in its stalk to directly contact the motor domain. This interaction prevents KIF17 from moving on microtubules. The cargo binding tail does not play an inhibitory role, but is nevertheless important in regulation as it functions to load KIF17 into cilia. KIF1A (Kinesin-3) is also inactive in the absence of cargo due to inhibitory regions that include the FHA domain and two adjacent coiled-coils. My work further demonstrates that KIF1A motors undergo processive motility as dimeric rather than monomeric motors using their neck coil to dimerize. Kinesin motors are relieved of their autoinhibition by cargo binding. However, as there are many different cargoes that require transport by the same kinesin motor, I have investigated whether specific cargo proteins compete or cooperate with each other for transport. My work shows that while some cargos are transported independently, other cargos such as JIP1 and JIP3 do form cooperative complexes with a single motor that are essential for efficient transport. The JIP1/ JIP3/Kinesin-1 complex is formed due to an interaction between JIP1 and JIP3 and distinct binding sites on Kinesin-1 for each cargo. Once cargo is loaded, kinesin motors have to transport them to their correct subcellular destinations. I have investigated whether Kinesin-1’s preference for certain types of modified microtubules can direct the polarized sorting of cargo into the axonal compartment of neuronal cells. My results indicate that several types of microtubule post-translational modifications additively or synergistically mediate Kinesin-1’s preference for axonal microtubules and its sorting mechanism. Overall, my work contributes to our understanding of how kinesin motors are activated and attached to their cargos in order to transport them to specific subcellular destinations.