Neuronal Substrates Required for Synapse Maintenance

Sreeganga Chandra, Program in Cellular Neuroscience, Neurodegeneration and Repair, Department of Neurology, Yale University

Synapses must be maintained throughout life to provide normal brain functions and behavior. Synapse maintenance is not passive, but an essential and actively regulated process in the adult brain. Drugs of abuse have been shown to induce both structural and functional changes in synapses, possibly by altering synapse maintenance pathways. Recently, several molecules important for maintaining synapses have been identified. Cysteine String Protein a (CSPa) and a-synuclein, presynaptic proteins that are widely expressed in the brain, are two such proteins. CSPa is a co-chaperone that assembles with Hsc70 and the tetratricopeptide-repeat protein SGT to form a chaperone complex on synaptic vesicles. Genetic studies in mice have shown that CSPa and a-synuclein cooperate in the maintenance of synapses.

Our long term goals are to define the pathways involved in synapse maintenance and to determine how they are altered in neurological disorders or upon addiction. In this proposal, we seek to identify the substrates of the CSPa chaperone complex by a proteomic comparison of synaptic fractions derived from wild type and CSPa knockout brains. We have opted to use proteomic approaches as the low affinity of the CSPa chaperone complex precludes conventional biochemical purifications. For this purpose, we will employ DIGE in concert with MALDI-MS/MS as well as iTRAQ. We will also identify those substrates that are restored by overexpression of a-synuclein using similar methods. Validated substrates will be tested in dissociated cultures for their roles in synapse maintenance and synaptic plasticity.

The diverse proteomic methodology and expertise of the NIDA Neuroproteomic Center will greatly aid us in addressing our specific aims. Through the planned experiments, we expect to identify key proteins that participate in synapse maintenance. This is a first and necessary step to understand the mechanisms underlying synapse maintenance and their contribution to alterations in neural circuits upon drug use.