Protein SUMOylation is a Sex-Specific Regulator of Fear Memory Formation in the Amygdala

SUMOylation is a type of post-translational protein modification similar to ubiquitination and it involves the covalent attachment of a small ubiquitin-like modifier (SUMO) protein to the lysine residue of a target substrate. While there is strong evidence for the role of protein ubiquitination in the formation of fear-based memories, few studies have been conducted examining the role that SUMOylation plays in this same process. The amygdala is the main site of storage for emotional memories and there is strong evidence that protein ubiquitination is critical for fear memory formation in this region. However, it has not previously been studied whether protein SUMOylation in the amygdala is also involved in fear memory formation. Additionally, although there is evidence to support sex differences in ubiquitin signaling during fear memory formation in the amygdala, whether males and females differ in their need for protein SUMOylation during fear memory formation has not been investigated. We have found significant sex differences in protein SUMOylation in the amygdala both at baseline (rest) and during fear memory formation. Western blot analysis revealed higher resting levels of SUMOylated proteins in females when compared to males, though both sexes showed global increases following fear conditioning. A SUMOylation-specific proteomic analysis discovered that only females had increased protein targeting with SUMO following fear conditioning, with four proteins being identified that gained SUMOylation modifications, the main target being a heat shock protein. One heat shock protein in males was identified as having lower SUMOylation levels following fear conditioning. This suggests sex differences in the interaction and targeting of proteins by SUMOylation following fear conditioning. We also inhibited the function of the only E2 conjugase for SUMOylation, Ube2i, via siRNA in the amygdala and found impaired fear memory in males but enhanced fear memory in females, though the latter only occurred under high siRNA concentrations. Interestingly, western blot analysis revealed that knockdown of Ube2i caused an increase in protein SUMOylation levels in females but a decrease in males, indicating that compensation is likely occurring in females. This suggests that in females, protein SUMOylation may be critical for basal cellular functioning, which precludes us from directly determining its role in fear memory formation. Collectively, these data reveal a novel, sex-specific role for protein SUMOylation in the amygdala during fear memory formation and expand our understanding of how ubiquitin-like signaling regulates memory formation.