Critical roles for the M3-S2 transduction linker domain in kainate receptor assembly and postassembly trafficking.
Kainate receptors (KARs) are neuronal proteins that exhibit a highly polarized distribution in the mammalian CNS. Assembly, intracellular trafficking, and synaptic targeting of KARs and other ionotropic glutamate receptors are processes controlled, in part, by various determinants within the constituent subunit proteins themselves. Here, we demonstrate that the linker region between the M3 and S2 domains, which in current structural models is thought to transduce ligand-binding energy into channel opening, additionally has an essential role in receptor biogenesis. Our results show that this gating-associated domain is engaged at two distinct critical stages of KAR biogenesis: first, during the transition from dimeric to tetrameric assembly states and, second, at a postassembly trafficking checkpoint within the endoplasmic reticulum. Alteration of a basic residue, arginine 663, altered the desensitization properties of the GluR6 kainate receptor in response to glutamate application, and these changes were weakly correlated with intracellular retention of the mutant receptors. Elimination of the positive charge also significantly attenuated oligomerization and stability of the intracellular subunit protein. Furthermore, charge swapping with an adjacent residue, glutamate 662, normalized the receptor physiological behavior and reversed the deficits in assembly and degradation, but only partially restored plasma membrane expression of the receptors. These results reveal a new role for this linker domain in glutamate receptor biogenesis and contribute to understanding the cellular controls of receptor assembly and trafficking, which will be important for relating receptor stoichiometry to their neuronal targeting and function.
Vivithanaporn
Pornpun
P
Department of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, Texas 77555, USA.
Lash
Laura Leanne
LL
Marszalec
William
W
Swanson
Geoffrey T
GT
D000818:Animals; D002462:Cell Membrane / Q000502:physiology; D006624:Hippocampus / Q000502:physiology*; D009154:Mutation; D009474:Neurons / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018092:Kainic Acid Receptors / Q000096:biosynthesis* / Q000235:genetics; D000097806:GluK2 Kainate Receptor
D018092:Kainic Acid Receptors; D000097806:GluK2 Kainate Receptor