SUMMARY The actin-binding protein adducin was recently identified as a component of the neuronal subcortical cytoskeleton. Here we analyzed mice lacking adducin to uncover the function of this protein in actin rings. α-adducin knockout mice presented progressive axon enlargement in the spinal cord, optic and sciatic nerve, followed by axon degeneration and loss. Using stimulated emission depletion super-resolution microscopy, we show that a periodic subcortical actin cytoskeleton is assembled in every neuron type inspected including retinal ganglion cells and dorsal root ganglia neurons. In neurons devoid of adducin, the actin ring diameter increased, although the inter-ring periodicity was maintained. In vitro, the actin ring diameter adjusted as axons grew suggesting the lattice is dynamic. Our data support a model in which adducin activity is not essential for actin ring assembly and periodicity, but is necessary to control the diameter of both actin rings and axons, and actin filament growth within rings.

Neurons have a membrane periodic skeleton (MPS) composed of actin rings interconnected by spectrin. Here, combining chemical and genetic gain- and loss-of-function assays, we show that in rat hippocampal neurons the MPS is an actomyosin network that controls axonal expansion and contraction. Using super-resolution microscopy, we analyzed the localization of axonal non-muscle myosin II (NMII). We show that active NMII light chains are colocalized with actin rings and organized in a circular periodic manner throughout the axon shaft. In contrast, NMII heavy chains are mostly positioned along the longitudinal axonal axis, being able to crosslink adjacent rings. NMII filaments can play contractile or scaffolding roles determined by their position relative to actin rings and activation state. We also show that MPS destabilization through NMII inactivation affects axonal electrophysiology, increasing action potential conduction velocity. In summary, our findings open new perspectives on axon diameter regulation, with important implications in neuronal biology.

In the adult nervous system axon caliber varies widely amongst different tracts. When considering a given axon, its diameter can further fluctuate in space and time, according to processes including the distribution of organelles and activity-dependent mechanisms. In addition, evidence is emerging supporting that in axons circumferential tension/contractility is present. Axonal diameter is generically regarded as being regulated by neurofilaments. When neurofilaments are absent or low, microtubule-dependent mechanisms can also contribute to the regulation of axon caliber. Despite this knowledge, the fine-tune mechanisms controlling diameter and circumferential tension throughout the lifetime of an axon, remain largely elusive. Recent data supports the role of the actin-spectrin-based membrane periodic skeleton and of non-muscle myosin II in the control of axon diameter. However, the cytoskeletal arrangement that underlies circumferential axonal contraction and expansion is still to be discovered. Here, we discuss in a critical viewpoint the existing knowledge on the regulation of axon diameter, with a specific focus on the possible role played by the axonal actin cytoskeleton.