Abstract
Polymerization and organization of actin filaments into complex superstructures is indispensable for structure and function of neuronal networks. We here report that knock down of the F-actin-binding protein Abp1, which is important for endocytosis and synaptic organization, results in changes in axon development virtually identical to Arp2/3 complex inhibition, i.e., a selective increase of axon length. Our in vitro and in vivo experiments demonstrate that Abp1 interacts directly with N-WASP, an activator of the Arp2/3 complex, and releases the autoinhibition of N-WASP in cooperation with Cdc42 and thereby promotes N-WASP-triggered Arp2/3 complex-mediated actin polymerization. In line with our mechanistical studies and the colocalization of Abp1, N-WASP and Arp2/3 at sites of actin polymerization in neurons, we reveal an essential role of Abp1 and its cooperativity with Cdc42 in N-WASP-induced rearrangements of the neuronal cytoskeleton. We furthermore show that introduction of N-WASP mutants lacking the ability to bind Abp1 or Cdc42, Arp2/3 complex inhibition, Abp1 knock down, N-WASP knock down and Arp3 knock down, all cause identical neuromorphological phenotypes. Our data thus strongly suggest that these proteins and their complex formation are important for cytoskeletal processes underlying neuronal network formation.
Highlights
The organization and dynamics of the cortical actin cytoskeleton play important roles in cell migration, establishment and changes of cell morphology and adhesion and in cellular uptake processes, such as phagocytosis, macropinocytosis and receptormediated endocytosis [1,2,3,4,5] – processes, which are indispensable for individual cells, the formations of larger cellular networks and organogenesis.The importance of actin filament polymerization and dynamics in neuronal cells has been primarily investigated during neuronal development
We here report that the mammalian F-actin-binding protein Abp1 plays an indispensable role in Arp2/3 complex-mediated actin polymerization controlled by the Arp2/3 complex activator N-WASP
We demonstrate that Abp1 directly interacts with N-WASP and that Abp1/N-WASP complexes exist both in brain extracts and in intact cells
Summary
The organization and dynamics of the cortical actin cytoskeleton play important roles in cell migration, establishment and changes of cell morphology and adhesion and in cellular uptake processes, such as phagocytosis, macropinocytosis and receptormediated endocytosis [1,2,3,4,5] – processes, which are indispensable for individual cells, the formations of larger cellular networks and organogenesis.The importance of actin filament polymerization and dynamics in neuronal cells has been primarily investigated during neuronal development. The organization and dynamics of the cortical actin cytoskeleton play important roles in cell migration, establishment and changes of cell morphology and adhesion and in cellular uptake processes, such as phagocytosis, macropinocytosis and receptormediated endocytosis [1,2,3,4,5] – processes, which are indispensable for individual cells, the formations of larger cellular networks and organogenesis. Since the Arp2/3 complex is involved in diverse actin cytoskeletal functions, specificity for individual processes and morphological features can only be brought about by distinct regulatory pathways and by further accessory components of this core actin nucleation machinery. The in vivo evidence for Arp2/3-complexdependent functions of a majority of these factors is sparse, their diversity may offer opportunities for cooperative regulations and may, in time and space, define the linkage of Arp2/3 complexmediated actin nucleation machinery to different cellular processes relying on actin polymerization
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