Characterization of a metabotropic glutamate receptor type 5-green fluorescent protein chimera (mGluR5-GFP): pharmacology, surface expression, and differential effects of Homer-1a and Homer-1c

Mol Cell Neurosci. 2001 Sep;18(3):296-306. doi: 10.1006/mcne.2001.1031.

Abstract

Metabotropic glutamate receptor 5 (mGluR5) can modulate synaptic transmission by increasing intracellular Ca2+ and it plays a role in several forms of synaptic plasticity. We have constructed a fusion of human mGluR5 and green fluorescent protein (mGluR5-GFP). Expression of mGluR5-GFP in clonal cell lines yielded a functional fluorescent receptor with pharmacological profiles similar to wild-type mGluR5. mGluR5-GFP coimmunoprecipitated with Homer-1c, indicating that addition of GFP to the C-terminal did not prevent Homer binding. Coexpression of wild-type mGluR5 or mGluR5-GFP with Homer 1c, but not Homer-1a, resulted in reduced receptor surface localization and the formation of intracellular clusters. Neither Homer-1a nor Homer-1c had any effect on mGluR1 or mGluR1-GFP distribution. mGluR5-GFP expressed alone or in combination with Homer-1a formed dimers in HEK cells. Coexpression with Homer-1c, however, prevented mGluR5-GFP dimerization. Neither Homer altered the agonist profiles of mGluR5 or mGluR5-GFP. These data indicate that the functional expression of mGluR5 is regulated by Homer-1c and demonstrate that mGluR5-GFP provides a useful tool to study the molecular pharmacology and cell biology of mGluRs in real-time.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CHO Cells / drug effects
  • CHO Cells / metabolism
  • Carrier Proteins / biosynthesis
  • Carrier Proteins / physiology*
  • Cricetinae
  • Dimerization
  • Dose-Response Relationship, Drug
  • Excitatory Amino Acid Antagonists / pharmacology
  • Green Fluorescent Proteins
  • Homer Scaffolding Proteins
  • Humans
  • Immunoblotting
  • Luminescent Proteins / genetics*
  • Neuropeptides / biosynthesis
  • Neuropeptides / physiology*
  • Precipitin Tests
  • Pyridines / pharmacology
  • Receptor, Metabotropic Glutamate 5
  • Receptors, Metabotropic Glutamate / antagonists & inhibitors
  • Receptors, Metabotropic Glutamate / biosynthesis
  • Receptors, Metabotropic Glutamate / genetics*
  • Receptors, Metabotropic Glutamate / metabolism*
  • Recombinant Fusion Proteins / antagonists & inhibitors
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / genetics*
  • Recombinant Fusion Proteins / metabolism*
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Transfection / methods

Substances

  • Carrier Proteins
  • Excitatory Amino Acid Antagonists
  • GRM5 protein, human
  • Homer Scaffolding Proteins
  • Luminescent Proteins
  • Neuropeptides
  • Pyridines
  • Receptor, Metabotropic Glutamate 5
  • Receptors, Metabotropic Glutamate
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins
  • 6-methyl-2-(phenylethynyl)pyridine