Interaction of serotonin 5-hydroxytryptamine type 2C receptors with PDZ10 of the multi-PDZ domain protein MUPP1

By using the yeast two-hybrid system, we previously isolated a cDNA clone encoding a novel member of the multivalent PDZ protein family called MUPP1 containing 13 PDZ domains. Here we report that the C terminus of the Fi-hydroxytryptamine type 2C (5-HT2C) receptor selectively interacts with the 10th PDZ domain of MUPP1. Mutations in the extreme C terminal SSV sequence of the 5-HT2C receptor confirmed that the SXV motif is critical for the interaction, Co-immunoprecipitations of MUPP1 and 5-HT2C receptors from transfected COS-7 cells and from rat choroid plexus verified this interaction in vivo. Immunocytochemistry revealed an SXV motif-dependent co-clustering of both proteins in transfected COS-7 cells as well as a colocalization in rat choroid plexus. A 5-HT2C receptor-dependent unmasking of a C-terminal vesicular stomatitis virus epitope of MUPP1 suggests that the interaction triggers a conformational change within the MUPP1 protein. Moreover, 5-HT2A and 5-HT2B, sharing the C-terminal EX(V/I)SXV sequence with 5-HT2C receptors, also bind MUPP1 PDZ domains in vitro. The highest MUPP1 mRNA levels were found in all cerebral cortical layers, the hippocampus, the granular layer of the dentate gyrus, as well as the choroid plexus, where 5-HT2C receptors are highly enriched. We propose that MUPP1 may serve as a multivalent scaffold protein that selectively assembles and targets signaling complexes.