Engineering Approaches to Biomolecular Motors

Engineering Approaches to Biomolecular Motors: From in vitro to in vivo Poster Abstracts

5-POS

Board 5

Cargo Rigidity Affects the Sensitivity of Dynein Ensembles to Individual Motor Pausing

Amalia Driller-Colangelo

, Jessica Morgan, Karen Chau, Nathan D. Derr.

Smith College, Northampton, MA, USA.

Cytoplasmic dynein is a minus-end directed microtubule-based motor protein that drives

intracellular cargo transport in eukaryotic cells. While many intracellular cargos are propelled by

small groups of dynein motors, many of the biophysical mechanisms that govern ensemble

motility remain unknown. We have designed a programmable DNA origami synthetic cargo

“chassis” that allows us to control the number of dynein motors in the ensemble and vary the

rigidity of the cargo chassis itself. On this chassis, motors within an ensemble are conjugated

together through variable length cargo “linkers” comprised of parallel segments of either single-

or double-stranded DNA. These regions determine the number of independent steps each motor

can take before exerting forces on the other motors within the ensemble. This design enables

investigation of how motor steps and pauses are “communicated” through the cargo structure and

how they affect the emergent behavior of the ensemble. Using TIRF microscopy, we have

observed dynein ensembles transporting these cargo chassis along microtubules in vitro. We find

that ensembles of dynein on flexible cargos move faster as more motors are added, whereas

ensembles on rigid cargos move slower as more motors are added. Using the slowly-

hydrolyzable ATP analog ATP-gamma-S, we have observed that ensembles connected through

flexible cargos are less sensitive to individual motor pausing. Our results suggest that the role of

cargo rigidity in the communication of motor pausing plays an important role in determining the

collective motility of dynein motor ensembles. The ability for cargos propelled by dynein

ensembles to maintain their motility despite the pausing of individual motors may allow the

cargo to maintain productive transport regardless of pauses induced by single motors

encountering obstacles on the microtubule.