Organelle Transport along Microtubules in Xenopus Melanophores: Evidence for Cooperation between Multiple Motors

Abstract

Xenopus melanophores have pigment organelles or melanosomes which, in response to hormones, disperse in the cytoplasm or aggregate in the perinuclear region. Melanosomes are transported by microtubule motors, kinesin-2 and cytoplasmic dynein, and an actin motor, myosin-V. We explored the regulation of melanosome transport along microtubules in vivo by using a new fast-tracking routine, which determines the melanosome position every 10 ms with 2-nm precision. The velocity distribution of melanosomes transported by cytoplasmic dynein or kinesin-2 under conditions of aggregation and dispersion presented several peaks and could not be fit with a single Gaussian function. We postulated that the melanosome velocity depends linearly on the number of active motors. According to this model, one to three dynein molecules transport each melanosome in the minus-end direction. The transport in the plus-end direction is mainly driven by one to two copies of kinesin-2. The number of dyneins transporting a melanosome increases during aggregation, whereas the number of active kinesin-2 stays the same during aggregation and dispersion. Thus, the number of active dynein molecules regulates the net direction of melanosome transport. The model also shows that multiple motors of the same polarity cooperate during the melanosome transport, whereas motors of opposite polarity do not compete.

Powering membrane traffic in endocytosis and recycling

“…Dispersion and aggregation are driven by kinesin-2 and cytoplasmic dynein, respectively (see figure). Novel precision measurements indicate that 1 or 2 copies of kinesin-2 and 1-3 dyneins can move a melanosome 84 . Kinesin-2 and dynein compete for the same binding site on p150Glued of dynactin 143 (arrows in the figure) and also interact with the regulator of melanophore activity, protein kinase A (not shown) 144 .…”

“…A curious phenomenon that is frequently observed in various cell systems is a block of movement in both directions when one of the motors (for example, cytoplasmic dynein) is downregulated 81,82 . This could be explained by the close association of both classes of motor on their cargoes, which conceivably can occur either by direct interaction 83 or coordination on the membrane 76,84 (see below and BOX 3). A potentially new twist to the story of bidirectional transport is the recent finding that purified dynein-dynactin complexes can move bidirectionally on microtubules in vitro 85 .…”

On the use of in vivo cargo velocity as a biophysical marker

“…To obtain the form of the in vivo FVCs of molecular motors we calculated the relative load per motor (Supplemental Table 1) using data from three different reports [3,4,22]. For neuronal vesicle transport velocities were normalized on a per-run basis by the authors.…”

“…Moving cargos are typically driven by multiple motors [1][2][3][4]. Much of this transport is bi-directional [5,6], including mRNA particles [7], mitochondria [2,8], virus particles [9,10], neuronal vesicles [11], etc.…”

“…Three studies [3,4,22] suggest that cytosolic viscous drag experienced by moving cargos is the dominant contributor to load opposing the motion of the cargos, and of sufficient magnitude to significantly slow down the motors. If so, a cargo could only speed up due to an increase in propulsion force.…”