Characterization of neuronal synaptopodin reveals a myosin V-dependent mechanism of synaptopodin clustering at the post-synaptic sites

The spine apparatus (SA) is an endoplasmic reticulum-related organelle which is present in a subset of dendritic spines in cortical and pyramidal neurons. The synaptopodin protein localizes between the stacks of the spine apparatus and is essential for the formation of this unique organelle. Although several studies have demonstrated the significance of the SA and synaptopodin in calcium homeostasis and plasticity of dendritic spines, it is still unclear what factors contribute to its stability at the synapse and whether the SA is locally formed or it is actively delivered to the spines. In this study we show that synaptopodin clusters are stable at their locations. We found no evidence of active microtubule-based transport for synaptopodin. Instead new clusters were emerging in the spines, which we interpret as the SA being assembled on-site. Furthermore, using super-resolution microscopy we show a tight association of synaptopodin with actin filaments. We identify the actin-based motor proteins myosin V and VI as novel interaction partners of synaptopodin and demonstrate that myosin V is important for the formation and/or maintenance of the SA.