Function of the first intracellular loop for human D1 and D5 dopaminergic receptors

Abstract

The human D1‐class dopamine receptors (hD1R and hD5R) are positively linked to adenylyl cyclase (AC) activation and play critical physiological roles in the brain. To study the function of their first intracellular loop (IL1), we have designed human D1‐ class mutants (hD1‐PD1 and hD5‐PD1) whereby all serines and threonines of IL1 have been replaced by alanines and valines, respectively. Both PD1 mutants exhibited a loss of dopamine affinity but unaltered binding properties for inverse agonists. Furthermore, constitutive activity of PD1 mutants was abolished, implicating that serines and threonines of IL1 are critical for the transition of D1‐class receptors from inactive (R) to active (R*) states in the absence of activating ligands. Dopamine potencies for hD1‐PD1 and hD5‐PD1 were drastically reduced compared to those of parent receptors. Interestingly, despite such warranted uncoupling from the G protein, both PD1 mutants displayed a higher dopamine‐mediated maximal activation of AC in relation to wild‐types. We speculate that this observation is attributed to an impairment in dopamine‐mediated internalization by PD1 mutants. Taken together, our study has demonstrated that IL1 can modulate ligand binding, G protein‐coupling, and possibly receptor trafficking, poising this region as an important structural determinant for D1‐class receptors. This research was funded by NSERC (grant #203694–2010).