PICK1 Interacts with and Regulates PKC Phosphorylation of mGLUR7

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Abstract
The G-protein-coupled metabotropic glutamate receptor subtype 7a (mGluR7a) is a member of group III metabotropic glutamate receptors that plays an important role as a presynaptic receptor in regulating transmitter release at glutamatergic synapses. Here we report that the protein interacting with C-kinase (PICK1) binds to the C terminus (ct) of mGluR7a. In the yeast two-hybrid system, the extreme ct of mGluR7a was shown to interact with the PSD-95/Discs large/ZO-1 (PDZ) domain of PICK1. Pull-down assays indicated that PICK1 was retained by a glutathione S-transferase fusion of ct-mGluR7a. Furthermore, recombinant and native PICK1/mGluR7a complexes were coimmunoprecipitated from COS-7 cells and rat brain tissue, respectively. Confocal microscopy showed that both PICK1 and mGluR7a displayed synaptic colocalization in cultured hippocampal neurons. PICK1 has previously been shown to bind protein kinase C α-subunit (PKCα), and mGluR7a is known to be phosphorylated by PKC. We show a relationship between these three proteins using recombinant PICK1, mGluR7, and PKCα, where they were co-immunoprecipitated as a complex from COS-7 cells. In addition, PICK1 caused a reduction in PKCα-evoked phosphorylation of mGluR7a inin vitro phosphorylation assays. These results suggest a role for PICK1 in modulating PKCα-evoked phosphorylation of mGluR7a.