The multifunctional protein GC1q-R interacts specifically with the i3 loop arginine cluster of the vasopressin V2 receptor

In this study, we identified the multifunctional protein GC1q-R as a novel vasopressin V2 receptor (V2R) interacting protein. For this purpose, we have developed a proteomic approach combining pull-down assays using a cyclic peptide mimicking the third intracellular loop of V2R as a bait and mass spectrometry analyses of proteins isolated from either rat or human kidney tissues or the HEK 293 cell line. Co-immunoprecipitation of GC1q-R with the c-Myc-tagged h-V2R expressed in a HEK cell line confirmed the existence of a specific interaction between GC1q-R and the V2 receptor. Then, construction of a mutant receptor in i3 loop allowed us to identify the i3 loop arginine cluster of the vasopressin V2 receptor as the interacting determinant for GC1q-R interaction. Using purified receptor as a bait and recombinant (74–282) GC1q-R, we demonstrated a direct and specific interaction between these two proteins via the arginine cluster.