Venus fly trap domain of mGluR1 functions as a dominant negative against group I mGluR signaling.
Metabotropic glutamate receptors (mGluRs) form covalently linked homodimers and contain large, N-terminal extracellular ligand binding, "venus fly trap" (VFT) domains. These domains, when expressed separately, are secreted as disulfide linked dimers and can dimerize with full-length receptors. mGluR splice variants have been described that contain only this domain, but the consequences of their interaction on receptor signaling have not been explored. Here it is shown that an mGluR1 mutant containing only the VFT is retained on the cell surface when a full-length receptor is co-expressed. Further, when expressed in rat superior cervical ganglion (SCG) neurons and modulation of native calcium currents is used as an assay for receptor activity, the VFT acts as a dominant negative with respect to mGluR1 signaling. Although full-length mGluR1 and mGluR5 are not known to heterodimerize, the mGluR5 VFT partially occludes mGluR1 signaling and the mGluR1 VFT potently occludes mGluR5 signaling in SCG neurons. In addition, an mGluR1 point mutant, mGluR1 C140G, which cannot covalently dimerize, functions like the wild-type receptor when expressed alone. The C140G mutant is inhibited by the mGluR1 VFT construct but does not retain the mGluR1 VFT on the cell surface, suggesting that the loss of C140 renders the interaction reversible. Finally, a peptide designed to disrupt mGluR1 dimerization reduced signaling through the C140G mutant receptor, but only when applied intracellularly for several hours, indicating that loss of signaling requires disruption of dimerization prior to plasma membrane insertion.
Beqollari
Donald
D
Department of Pharmacology and Physiology, University of Rochester Medical Center, Rochester, New York 14642, USA.
Kammermeier
Paul J
PJ
D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D019281:Dimerization; D005110:Extracellular Space / Q000378:metabolism / Q000502:physiology; D008297:Male; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D017354:Point Mutation / Q000502:physiology; D011485:Protein Binding; D011487:Protein Conformation; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D064529:Receptor, Metabotropic Glutamate 5; D018091:Receptors, AMPA / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D018094:Receptors, Metabotropic Glutamate / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D017783:Superior Cervical Ganglion / Q000166:cytology / Q000502:physiology
D004247:DNA; D016271:Proto-Oncogene Proteins c-myc; D064529:Receptor, Metabotropic Glutamate 5; D018091:Receptors, AMPA; D018094:Receptors, Metabotropic Glutamate; C578937:Grm5 protein, rat