Plasticity of synapse nanostructure: dissecting the dynamic nanoscale organization of neuronal synapses

Activity-dependent changes in the molecular organization of synapses are broadly held to underlie the long-term changes in synaptic strength that are associated with learning and memory. In particular, the density and position of AMPA-type glutamate receptors (AMPARs) relative to the presynaptic release site have emerged as critical determinants of synaptic efficacy. Thus, identifying the mechanisms that control the positioning of AMPARs at synapses is critical for a fundamental understanding of synaptic transmission and plasticity. Recently, we used super-resolution microscopy to demonstrate that the spatial distribution of synaptic components is highly heterogeneous and that these molecular assemblies undergo structural reorganization in response to synaptic activity. Importantly, these studies demonstrated that not only the number of AMPARs, but also their subsynaptic localization with respect to the presynaptic release site is a critical determinant of synaptic strength. However, the molecular mechanisms that underlie the¬¬se structural adaptations and how these mechanisms contribute to plasticity-induced changes in synaptic efficacy remain unresolved. This is in large part due to the limited ability of conventional microscopy techniques to resolve the distribution of proteins in individual synapses, but also due to the lack of tools that efficiently and specifically label endogenous proteins. To overcome these technical hurdles, we developed CRISPR/Cas9-mediated genome editing tools that allow efficient and specific labelling of endogenous proteins with fluorescent tags that are compatible with super-resolution imaging techniques. Thus, these innovations allow us to resolve the distribution of key synaptic proteins at endogenous levels with nanoscale precision. In this project, we will combine these innovations with electrophysiological assays to 1) measure activity-dependent changes in distribution patterns of synaptic components, 2) determine the molecular interactions that underlie the structural and functional adaptation of synapses, and 3) identify trans-synaptic interactions that define the molecular organization of synapses to sustain a long-term increase in synaptic efficacy. Altogether, this project will dissect, at the molecular level, how neuronal synapses adapt to express the plasticity-induced changes in synaptic strength that underlie cognitive processes such as learning and memory.