Elizabeth A. Eipper, Department of Neuroscience, University of Connecticut Health Center
The technologies available to us through the Neuroproteomics Center will allow us to determine the phosphorylation state of Kalirin7 in the mouse striatum/nucleus accumbens under basal conditions and after acute and chronic exposure to cocaine. A multiple reaction monitoring method will be developed so that phosphorylation at all key sites can be determined simultaneously. Cdk5, whose role in addiction is studied by Center Investigator James Bibb, alters Kalirin7 function by phosphorylating a single site near its C-terminus. Methods for assessing the phosphorylation state of activated Kalirin7 and Kalirin7 that is engaged in interactions with the NR2B subunit of the NMDA receptor will be developed. Lfc, a RhoGEF found in dendritic spines but not concentrated at the PSD, is studied by Center Director Angus Nairn; comparisons of Kalirin7 and Lfc will be especially informative. The 8-plex iTraq capabilities developed in the Center will allow us to compare the composition of PSDs isolated from wildtype and Kal7KO mice under control conditions and after chronic exposure to cocaine. Kalirin7, with its nine spectrin-like repeats and PDZ-binding motif, is known to interact with multiple proteins; its absence may result in the loss of specific components of the PSD. The Kal7KO mice, with their heightened sensitivity to cocaine, should provide a valuable resource to Center Investigators studying synaptic maintenance, protein complexes that mediate synapse formation and NMDA Receptor complexes.