Myosin Va transport of liposomes in three-dimensional actin networks is modulated by actin filament density, position, and polarity

Abstract

Significance Intracellular transport of critical cellular components (e.g. vesicles, organelles, mRNA, chromosomes) is accomplished by myosin Va molecular motors along complicated 3D networks of actin filaments. Disruption of these transport processes leads to debilitating human disease (e.g. Griscelli syndrome), while rearrangement of the 3D actin cytoskeleton is a hallmark of malignant cancers. We found that the various modes of motion (stationary, diffusive-like, or directed) describing how teams of myosin Va transport 350-nm liposome cargos are determined by the 3D position and polarity of the actin filaments within the network that the myosin Va motors interact with. This study demonstrates that the 3D actin filament organization within the network can serve as a potent regulator of myosin Va motor-based intracellular transport. The cell’s dense 3D actin filament network presents numerous challenges to vesicular transport by teams of myosin Va (MyoVa) molecular motors. These teams must navigate their cargo through diverse actin structures ranging from Arp2/3-branched lamellipodial networks to the dense, unbranched cortical networks. To define how actin filament network organization affects MyoVa cargo transport, we created two different 3D actin networks in vitro. One network was comprised of randomly oriented, unbranched actin filaments; the other was comprised of Arp2/3-branched actin filaments, which effectively polarized the network by aligning the actin filament plus-ends. Within both networks, we defined each actin filament’s 3D spatial position using superresolution stochastic optical reconstruction microscopy (STORM) and its polarity by observing the movement of single fluorescent reporter MyoVa. We then characterized the 3D trajectories of fluorescent, 350-nm fluid-like liposomes transported by MyoVa teams (∼10 motors) moving within each of the two networks. Compared with the unbranched network, we observed more liposomes with directed and fewer with stationary motion on the Arp2/3-branched network. This suggests that the modes of liposome transport by MyoVa motors are influenced by changes in the local actin filament polarity alignment within the network. This mechanism was supported by an in silico 3D model that provides a broader platform to understand how cellular regulation of the actin cytoskeletal architecture may fine tune MyoVa-based intracellular cargo transport.