A glycosylation-specific interaction between the Sigma-1 and Neuorkinin-1 receptors (659.13)

The putative chaperone protein Sigma-1 Receptor (S1R) has been implicated in chronic pain via pharmacological and gene knockout studies, however, how S1R affects pain is not known. Activation of the GPCR Neurokinin-1 Receptor (NK1R) is implicated in pain signaling. Herein we demonstrate a novel robust interaction between S1R (FLAG-tagged) and NK1R (HA-tagged) using co-immunoprecipitation (co-IP) in transiently transfected HEK293 cells. NK1R are glycoproteins with several mobility-shifted immunoreactive bands present on a Western blot. Co-IP demonstrates specific interaction between S1R and only the non-glycosylated NK1R. This specific interaction was confirmed by tunicamycin-treatment to eliminate the glycosylation and by expression of a glycosylation-deficient NQ-NK1R mutant. Additionally, we assay a panel of GPCRs for S1R interaction, (including the NK2R, NK3R, MOR, DA1, and MRGR), and demonstrate that all of the interactions are restricted to the non-glycosylated GPCR. We conclude that S1R acts as a pr...