Cytoskeletal self-organization in neuromorphogenesis

Abstract

Self-organization of dynamic microtubules via interactions with associated motors plays a critical role in spindle formation. The microtubule-based mechanisms underlying other aspects of cellular morphogenesis, such as the formation and development of protrusions from neuronal cells is less well understood. In a recent study, we investigated the molecular mechanism that underlies the massive reorganization of microtubules induced in non-neuronal cells by expression of the neuronal microtubule stabilizer MAP2c. In that study we directly observed cortical dynein complexes and how they affect the dynamic behavior of motile microtubules in living cells. We found that stationary dynein complexes transiently associate with motile microtubules near the cell cortex and that their rapid turnover facilitates efficient microtubule transport. Here, we discuss our findings in the larger context of cellular morphogenesis with specific focus on self-organizing principles from which cellular shape patterns such as the thin protrusions of neurons can emerge.

Keywords: cellular morphogenesis; cortical dynein; cytoplasmic dynein; microtubules; neurite; neuromorphogenesis; self-organization.

Figure 1. Cytoskeletal organization of neurons before and after neurite formation. Neurites are thin protrusions that originate from neuronal cell bodies. They are characterized by a microtubule-rich shaft and tipped by an F-actin-rich growth cone. Microtubules: red; F-actin: green; red and green arrows: forces based on microtubules and the F-actin cytoskeleton, respectively. Microtubule plus tips are indicated by ⊕ symbols.

Figure 2. Direct observation of microtubule pushing by cortical dynein in living cells. (A) Images of COS7 cells transfected with the neuronal microtubule stabilizer MAP2c obtained shortly after nocodazole washout. In those conditions, short microtubules formed that were transported directionally within a distance up to ~200nm from the plasma membrane as observed by evanescent wave illumination in total internal refelction fluorescence (TIRF) microscopy. (B) Individual frames from video-microscopic observation of the boxed region in A. (C) Simultaneous observation of MAP2c-decorated short microtubules (left) and dynein heavy chain subunits (Dync1h1, middle) via TIRF microscopy. As expected for a direct, physical interaction between those components, the fluorescence signal derived from MAP2c-decorated microtubules is strongest at the localization of cortical dynein, suggesting that this microtubule region is closest to the plasma membrane and therefore excited most strongly by the evanescent wave of the TIRF microscope. (D) Schematic of the dynein-mediated microtubule pushing mechanism. This image was taken from (http://www.molbiolcell.org/content/25/1/95).

Figure 3. Model for MAP2c and dynein-mediated self-organization of microtubules. (A) Simulations based on mathematical modeling of cortical dynein mediated microtubule transport closely mimic experimental observations after nocodazole washout in the presence of MAP2c. Both the overall redistribution (shown here) and saltatory movements of individual microtubules are observed in those simulations. The white arrow points to microtubules radiating from a microtubule organizing center that was not included in simulations. (B) Shortly after washout of nocodaole, small microtubules with random initial orientation nucleate in the cytosol. Those microtubules are transported directionally with leading plus ends via cortical dynein complexes until they encounter an obstacle, which ultimately will be the plasma membrane. This will orient microtubule plus ends to point toward the cell periphery. Microtubules will then push against the outer edge of the cell, which can induce a small convex cell protrusion. (C) Enlarged region marked in B. In a process related to stigmergy, the convex cell protrusion can self-amplify by collecting and trapping more microtubules, which push further against the cell periphery. (D) Distinct modes of interaction between microtubules and cortical dynein. Right: The “end-on” dynein-microtubule interaction produces pulling forces that are integrated to pull the MTOC toward regions of higher cortical dynein density. At homogenous densities, pulling forces can lead to MTOC centering. Left: The “side-on” dynein-microtubule interaction produces pushing forces that are integrated to push microtubules toward the cell periphery where they can stimulate cell protrusions via a “clutch”-like mechanism. Panel A was taken from (http://www.molbiolcell.org/content/25/1/95).