Remote Control of Diverse Cytoskeletal Motors using Light-Activated Gearshifting

Abstract

Cytoskeletal motors perform critical force generation and transport functions in eukaryotic cells. Protein engineering has been used to modify cytoskeletal motors for dynamic control of activity and directionality, providing direct tests of structure-function relationships and potential tools for controlling cellular processes or for harnessing molecular transport in artificial systems. We have previously1 created myosin motors that can be signaled to switch directions in response to changes in [Ca2+]. Light is a more versatile control signal2 because it can be precisely modulated in space and time, and is generally orthogonal to cellular signaling. Here we report the design and characterization of a panel of cytoskeletal motors that reversibly change gears - speed up, slow down, or switch directions - when exposed to blue light. Our structural designs incorporate a photoactive protein domain to enable light-dependent conformational changes in an engineered lever arm. We have used in vitro motility assays to confirm robust spatiotemporal control over motor function and to characterize the kinetics of optical gearshifting. Our modular approach has yielded controllable motors for both actin-based and microtubule-based transport. Genetically encoded light-responsive motors will expand the optogenetics toolkit, complementing precise perturbations of ion channels and intracellular signaling with spatiotemporal control of cytoskeletal transport and contractility.1. Chen, L., Nakamura, M., Schindler, T.D., Parker, D. & Bryant, Z. Engineering controllable bidirectional molecular motors based on myosin. Nature nanotechnology 7, 252-6 (2012).2. Walter, W.J. & Diez, S. Myosin shifts into reverse gear. Nature nanotechnology7, 213-4 (2012).