UC San Diego

Leucine-Rich Repeat Kinase 2 (LRRK2) is one of the most commonly mutated genes in familial Parkinson’s Disease (PD). This neurodegenerative disease affects over 10 million people worldwide, and defects in LRRK2 function have also been linked to the sporadic form of the disease. LRRK2 is a large (286 kDa) multi-domain, predominantly cytosolic, protein and has been shown to have roles in membrane trafficking. While a subset of LRRK2 is localized to membranes in cells, LRRK2 has also been shown to colocalize with microtubules and form helical filaments that wrap around the microtubules. Intriguingly, many PD-linked mutations demonstrate an enhanced association with microtubules when expressed in cells. In vitro, microtubule-associated LRRK2 can act as a roadblock for the molecular motor proteins dynein and kinesin. However, many questions remain about the impact this microtubule-associated LRRK2 has on intracellular transport and how these LRRK2 filaments form. In this dissertation, I present interdisciplinary work investigating the mechanisms by which LRRK2 filament formation occurs. Here, we report a cryo-electron microscopy structure of the catalytic half of LRRK2, containing its kinase, which is in a closed conformation, and GTPase domains, bound to microtubules. We also report a structure of the catalytic half of LRRK1, LRRK2’s closest human homolog, which, while structurally similar to LRRK2, is not linked to PD and does not interact with microtubules. Guided by these structures, we identify amino acids in LRRK2’s GTPase domain that mediate microtubule binding; mutating them disrupts microtubule binding in vitro and in cells without affecting LRRK2’s kinase activity. Overall, this work provides important insights into the basis of LRRK2 binding to microtubules and presents novel mutants that will enable future work to better probe the physiological roles of the LRRK2/microtubule interaction in the pathogenesis of PD. Our results have implications for the design of therapeutic LRRK2 kinase inhibitors.