Tug-of-war between dissimilar teams of microtubule motors regulates transport and fission of endosomes

Abstract

Intracellular transport is interspersed with frequent reversals in direction due to the presence of opposing kinesin and dynein motors on organelles that are carried as cargo. The cause and the mechanism of reversals are unknown, but are a key to understanding how cargos are delivered in a regulated manner to specific cellular locations. Unlike established single-motor biophysical assays, this problem requires understanding of the cooperative behavior of multiple interacting motors. Here we present measurements inside live Dictyostelium cells, in a cell extract and with purified motors to quantify such an ensemble function of motors. We show through precise motion analysis that reversals during endosome motion are caused by a tug-of-war between kinesin and dynein. Further, we use a combination of optical trap-based force measurements and Monte Carlo simulations to make the surprising discovery that endosome transport uses many (approximately four to eight) weak and detachment-prone dyneins in a tug-of-war against a single strong and tenacious kinesin. We elucidate how this clever choice of dissimilar motors and motor teams achieves net transport together with endosome fission, both of which are important in controlling the balance of endocytic sorting. To the best of our knowledge, this is a unique demonstration that dynein and kinesin function differently at the molecular level inside cells and of how this difference is used in a specific cellular process, namely endosome biogenesis. Our work may provide a platform to understand intracellular transport of a variety of organelles in terms of measurable quantities.asymmetric motor competition ͉ coordination of motors ͉ molecular motor dynein kinesin ͉ regulation of bidirectional motion ͉

Intracellular transport driven by cytoskeletal motors: General mechanisms and defects

“…As a result, the cargo will go back and forth, possibly performing large (compared to the typical motor step size) excursions in each direction [156]. An experimental observation in favour of this scenario was made on endosomes which were found to be elongated when pausing [351]. This could be a consequence of the pulling of opposite teams during tug-of-war episodes.…”

“…Detachment and diffusion periods could be longer than in vitro as they take place in a viscous environment [244]. However, estimations of the number of attached motors (around five of each type in [215], 1 or 2 kinesins against 4 to 8 dyneins in [351]) make the full detachment of the cargo quite unlikely. Competition between different types of motors, or obstacles could also explain these pauses.…”

“…Experimental evidence for the attachment of many motors is based on a number of observations, in particular based on force measurements [345,351,215,223]. Another argument is that, for several motor-cargo systems, there is evidence for bidirectional non diffusive motion [388].…”

Segmentation of 3D Trajectories Acquired by TSUNAMI Microscope: An Application to EGFR Trafficking

“…5) derived from phase III matched well with the typical motor protein-mediated transport speed of 0.5 to 2 mm/s (59-62). The two separate populations of V i in phase III might represent the two classes of runs (the short-slow run and the long-fast run) observed in active transport (85,86) (Fig. 4 F).…”

From the Cell Membrane to the Nucleus: Unearthing Transport Mechanisms for Dynein

“…Currently, the available experimental data does not show how the dynein itself is behaving at these times, as we only see the position of the endosome. The pauses and retrograde transport may be a result of the action of kinesins on the cargo, either in a tug-of-war scenario, dynein may occasionally fall off a microtubule, or simply become inhibited by a crowded cytoplasm [1,21,38,42,48]. In addition, non-radial movement, could also be a result of dynein being detached from the cargo or the microtubule.…”