Abstract
A range of cargo adaptor proteins are known to recruit cytoskeletal motors to distinct subcellular compartments. However, the structural impact of cargo recruitment on motor function is poorly understood. Here, we dissect the multimodal regulation of myosin VI activity through the cargo adaptor GAIP-interacting protein, C terminus (GIPC), whose overexpression with this motor in cancer enhances cell migration. Using a range of biophysical techniques, including motility assays, FRET-based conformational sensors, optical trapping, and DNA origami–based cargo scaffolds to probe the individual and ensemble properties of GIPC–myosin VI motility, we report that the GIPC myosin-interacting region (MIR) releases an autoinhibitory interaction within myosin VI. We show that the resulting conformational changes in the myosin lever arm, including the proximal tail domain, increase the flexibility of the adaptor–motor linkage, and that increased flexibility correlates with faster actomyosin association and dissociation rates. Taken together, the GIPC MIR–myosin VI interaction stimulates a twofold to threefold increase in ensemble cargo speed. Furthermore, the GIPC MIR–myosin VI ensembles yield similar cargo run lengths as forced processive myosin VI dimers. We conclude that the emergent behavior from these individual aspects of myosin regulation is the fast, processive, and smooth cargo transport on cellular actin networks. Our study delineates the multimodal regulation of myosin VI by the cargo adaptor GIPC, while highlighting linkage flexibility as a novel biophysical mechanism for modulating cellular cargo motility.
Highlights
IntroductionThe adaptor GIPC has been shown to recruit myosin VI to uncoated vesicles during endocytosis (1)
In addition to controlling motor density and nonspecific surface binding, the DNA nanotube assay geometry allows us to use saturating concentrations of GIPC myosin interacting region (MIR) (2 μM), allowing for a non-ambiguous interpretation of the effect of adapter binding on myosin VI speed
All three conditions had similar numbers of motile events per field of view (Figure 2C) and had similar run lengths (Figure 2D). These data highlight the dramatic effect of GIPC MIR on the ensemble speeds of myosin VI driven transport
Summary
The adaptor GIPC has been shown to recruit myosin VI to uncoated vesicles during endocytosis (1). By bridging the gap between the motor and proteins expressed on the cell surface, GIPC has been implicated as a key regulator of endocytosis for cellular signaling and adhesion molecules Further supporting this role, GIPC overexpression has been found in numerous cancers (3). Myosin VI exhibits load-dependent anchoring to actin filaments, demonstrating the potential for cargo-mediated changes in acto-myosin interaction kinetics (15) Despite this wealth of information on the structural features of myosin VI activity, it has mostly been obtained in the absence of cargo adaptors. The enhanced flexibility leads to faster motor-actin interaction kinetics by reducing the dwell time of the motor on actin filaments Taken together, these effects yield fast, smooth, processive movement of GIPC-recruited myosin VI ensembles on cellular actin networks
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
