Single molecule spectroscopy; Coordination of post-synaptic glutamate receptor signaling by the MAGUK family of scaffolds.

The Bowen Lab focuses on the membrane-associated guanylate kinase (MAGuK) family of scaffold proteins, which are key organizers of excitatory neurotransmission and are implicated in diseases such as neurodegeneration following stroke, autism, epilepsy and schizophrenia. Scaffolds determine the outcome of signal transduction by controlling the location of receptors and connecting them to downstream effectors. Studying scaffolds in vivo has been hampered by the complexity and redundancy of multiple isoforms present in the brain. Our approach is to reconstitute these cellular complexes on biological membranes to uncover how protein structure and posttranslational modifications, such as phosphorylation and palmitoylation, shape their scaffolding activity. By taking a reductionist approach, we can learn the rules that govern the spontaneous assembly of proteins within the postsynaptic density of excitatory neurons.

The proteins we study contain intrinsic disorder meaning they lack the fixed structures that give rise to activity in enzymes yet they mediate critical synaptic functions. The lack of a fixed structure challenges conventional approaches to structural biology so we use fluorescence microscopy to follow their transient interactions and refine molecular models for their dynamic structures. Harnessing imaging methodology for protein structure determination allows us to study previously intractable complexes. Our primary approach is to image at the level of single molecules, which allows us to follow the composition, structure and dynamics of an individual scaffold and its complexes with synaptic proteins. By combining the latest in single molecule methods with our working reconstitution of the postsynaptic membrane, we are working clarify the dynamic assembly of proteins that govern the response to glutamate neurotransmitters at the excitatory synapse.