Amino acid substitutions in the pore helix of GluR6 control inhibition by membrane fatty acids.
RNA editing at the Q/R site in the GluR5 and GluR6 subunits of neuronal kainate receptors regulates channel inhibition by lipid-derived modulators including the cis-unsaturated fatty acids arachidonic acid and docosahexaenoic acid. Kainate receptor channels in which all of the subunits are in the edited (R) form exhibit strong inhibition by these compounds, whereas wild-type receptors that include a glutamine (Q) at the Q/R site in one or more subunits are resistant to inhibition. In the present study, we have performed an arginine scan of residues in the pore loop of the GluR6(Q) subunit. Amino acids within the range from -19 to +7 of the Q/R site of GluR6(Q) were individually mutated to arginine and the mutant cDNAs were expressed as homomeric channels in HEK 293 cells. All but one of the single arginine substitution mutants yielded functional channels. Only weak inhibition, typical of wild-type GluR6(Q) channels, was observed for substitutions +1 to +6 downstream of the Q/R site. However, arginine substitution at several locations upstream of the Q/R site resulted in homomeric channels exhibiting strong inhibition by fatty acids, which is characteristic of homomeric GluR6(R) channels. Based on homology with the pore loop of potassium channels, locations at which R substitution induces susceptibility to fatty acid inhibition face away from the cytoplasm toward the M1 and M3 helices and surrounding lipids.
Department of Cell Biology and Physiology, Washington University Medical School, St. Louis, MO 63110, USA.
Timothy J
TJ
Wilding
Elisabeth
E
Fulling
Yun
Y
Zhou
James E
JE
Huettner
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000596:Amino Acids / Q000235:genetics; D000818:Animals; D001120:Arginine / Q000235:genetics; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003208:Concanavalin A / Q000494:pharmacology; D004281:Docosahexaenoic Acids / Q000494:pharmacology; D004594:Electrophysiology; D005227:Fatty Acids / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007608:Kainic Acid / Q000494:pharmacology; D008564:Membrane Potentials / Q000502:physiology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017393:RNA Editing; D051381:Rats; D018092:Kainic Acid Receptors / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D000097806:GluK2 Kainate Receptor
D000596:Amino Acids; D001120:Arginine; D003208:Concanavalin A; D004281:Docosahexaenoic Acids; D005227:Fatty Acids; D007608:Kainic Acid; D018092:Kainic Acid Receptors; D000097806:GluK2 Kainate Receptor