THE SCAFFOLDS OF MEMORY: SYNAPTIC ARCHITECTURE OF ENGRAM NEURONS

Persistent memory relies on stable synaptic modifications within engram cell networks. Although the medial prefrontal cortex (mPFC) plays a central role in the expression of remote (weeks- to months-old) memories, the molecular determinants that maintain synaptic stability in cortical engram cells remain largely unknown.

Here, we combined activity-dependent neuronal tagging (TRAP2-tdTomato mouse line) with biochemical isolation of synapses and fluorescence-activated synapse sorting, followed by unbiased nanoscale quantitative proteomics, to characterize the synaptic protein composition of synapses from engram and non-engram neurons in the mPFC four weeks after contextual fear conditioning.

Among the differentially expressed proteins were members of the SHANK family. SHANK proteins are postsynaptic scaffolding molecules that organize excitatory synapses by linking glutamate receptors to the actin cytoskeleton, and mutations in SHANK genes are strongly associated with synaptopathies and cognitive disorders. Their selective enrichment at synapses on engram neurons suggests a role in stabilizing long-lasting memory traces.

To investigate the functional relevance of SHANK in engram neurons, we employed engram-specific miRNA-mediated knockdown strategies in the mPFC and are assessing effects on synaptic connectivity and memory retrieval. Ongoing analyses examine how engram-specific SHANK depletion impacts excitatory synaptic transmission, spine morphology, and the long-term expression of remote fear memory.

Together, this work identifies upregulated synaptic scaffold proteins as a candidate molecular determinant of memory persistence and provides a framework to dissect mechanisms that stabilize cortical engram circuits.