At present, little is known regarding the mechanism of metabotropic glutamate receptor (mGluR) trafficking. To facilitate this characterization we inserted a haemagglutinin (HA) epitope tag in the extracellular N-terminal domain of the rat mGluR1a. In human embryonic kidney cells (HEK293), transiently transfected with HA-mGluR1a, the epitope-tagged receptor was primarily localized to the cell surface prior to agonist stimulation. Following stimulation with glutamate (10 µm; 30 min) the HA-mGluR1a underwent internalization to endosomes. Further quantification of receptor internalization was provided by ELISA experiments which showed rapid agonist-induced internalization of the HA-mGluR1a. To determine whether agonist-induced mGluR1a internalization is an arrestin- and dynamin-dependent process, cells were cotransfected with HA-mGluR1a and either of these dynamin-K44A or arrestin-2 (319–418). Expression of either dominant negative mutant constructs with receptor strongly inhibited glutamate-induced (10 µm; 30 min) HA-mGluR1a internalization. In addition, wild-type arrestin-2−green fluorescent protein (arrestin-2−GFP) or arrestin-3−GFP underwent agonist-induced translocation from cytosol to membrane in HEK293 cells coexpressing HA-mGluR1a. Taken together our observations demonstrate that agonist-induced internalization of mGluR1a is an arrestin- and dynamin-dependent process.
Mundell, S. J., Matharu, A-L., Pula, G., Roberts, P. J., & Kelly, E. (2001). Agonist-induced internalization of the metabotropic glutamate receptor 1a is arrestin- and dynamin-dependent. Journal of Neurochemistry, 78(3), 546-551. https://doi.org/10.1046/j.1471-4159.2001.00421.x