Measuring molecular motor forces in vivo: implications for tug-of-war models of bidirectional transport

Author :C. Leidel, R. A. Longoria, F. Marquez Gutierrez and G. T. Shubeita
Publisher :Biophysical J.
Volume :103
Number :3
Pages :492-500
Year :2012
Note :
Molecular motor proteins use the energy released from ATP hydrolysis to generate force and haul cargoes alongcytoskeletal ?laments. Thus, measuring the force motors generate amounts to directly probing their function. We report onoptical trapping methodology capable of making precise in vivo stall-force measurements of individual cargoes hauled by molecular motors in their native environment. Despite routine measurement of motor forces in vitro, performing and calibrating suchmeasurements in vivo has been challenging. We describe the methodology recently developed to overcome these dif?culties,and used to measure stall forces of both kinesin-1 and cytoplasmic dynein-driven lipid droplets in Drosophila embryos. Critically,by measuring the cargo dynamics in the optical trap, we ?nd that there is memory: it is more likely for a cargo to resume motion inthe same direction’rather than reverse direction’after the motors transporting it detach from the microtubule under the force ofthe optical trap. This suggests that only motors of one polarity are active on the cargo at any instant in time and is not consistentwith the tug-of-war models of bidirectional transport where both polarity motors can bind the microtubules at all times. We furtheruse the optical trap to measure in vivo the detachment rates from microtubules of kinesin-1 and dynein-driven lipid droplets.Unlike what is commonly assumed, we ?nd that dynein’s but not kinesin’s detachment time in vivo increases with opposingload. This suggests that dynein’s interaction with microtubules behaves like a catch bond.