Grant: Discovery Grants Program - Individual

The proper subcellular localization of proteins is fundamental for brain development. The global rules that dictate protein localization, however, remain poorly understood. Several lines of evidence suggest that protein SUMOylation (covalent modification of a small ubiquitin-like modifier [SUMO] to lysines) is not only important for protein localization but also for brain function. Specifically, Sumo2 (but not its paralogues Sumo1 and Sumo3) null mice die early on in development. Therefore, we theorized that identifying Sumo2 substrates would shed important light into brain function. To test this idea, we generated Sumo2 knockin mice which carry an HA epitope tag and pulled down HA-Sumo2-conjugates from the brains of mice, identifying the substrates via mass-spectrometry and comparing them to Sumo1 knockin mice (Tirard et al, 2012). When comparing the brain-resident Sumo2- and Sumo1-clients, we found only 28% overlapping proteins. This is supported by immunofluorescence staining showing drastically different subcellular localization of Sumo1 (strictly nuclear) vs. Sumo2 (broadly distributed). One of the most robust candidates picked up selectively in Sumo2 vs. Sumo1 pulldowns is the kinase CaMKII. This is exciting because CaMKII is a critical mediator of memory formation and is a tightly regulated protein. Its levels, phosphorylation and interactions with other proteins are known to mediate its activity. Nevertheless, the extent of its regulation is not completely understood. Given the critical role for protein SUMOylation in development, this mechanism may be a strong modulator in mediating CaMKII levels, localization, activity and ultimately its neural function. Moreover, our finding that Sumo2-conjugated CaMKII occurs endogenously, suggests a new tier of protein regulation which may be applied across numerous brain-resident protein substrates, thus allowing for an in-depth research program on the role of protein SUMOylation in the context of brain development. In the short term of this program, we will test how SUMOylation mediates CaMKII function. Specifically, we will test how these affect CaMKII protein levels, subcellular localization and on neural substrates of memory formation. Moreover, we will test whether neuronal activity modulates CaMKII SUMOylation in vitro. Lastly, we will spatially and temporally deconstruct CaMKII SUMOylation across different brain regions during development. Future studies will investigate how this modification affects mouse behaviours to ultimately understand the impact of SUMOylation on dictating protein function and the developmental fate of the brain. Long term, this program stands to benefit the neuroscience community as it provides an atlas of Sumo2-bound neural substrates and thus sheds important light into how this modification affects their function in the brain. Given the multiple substrates identified in this proteomics screen, several projects and HQP stand to benefit from its fruit.