Synapsin-1 and actin form ordered nanoscale assemblies.

Liquid-liquid phase separation is a vital and ubiquitous principle of subcellular organization. Several actin-binding proteins have recently been shown to undergo liquid-liquid phase separation, forming micron-sized droplets that assemble actin into distinct network shapes. However, our knowledge of how these phase-separated proteins organize actin filaments at the nanoscale is rather limited. Here, we seek to address this knowledge gap by investigating synapsin-1 condensates through a combination of computational simulations and nanoscale imaging experiments. Our results show that while synapsin-1 condensates by themselves lack any special structural organization, the addition of actin filaments results in the generation of a regularly organized actin scaffold within the condensates. The organized scaffold forms both when actin and synapsin-1 are added simultaneously and when actin is added after the formation of synapsin condensates. Dissolving the condensates, by synapsin-1 removal, leaves the actin scaffold largely unaffected in both simulations and experiments, implying that this scaffold should guide droplet reformation when synapsin-1 is added back. Taken together, our findings suggest that the actin-rich component of the condensate retains a structural memory that promotes and guides the reassembly of synapsin-actin condensates. This structural memory may be important in the long-term maintenance of synaptic function in vivo .