title: Channel-opening kinetic mechanism of wild-type GluK1 kainate receptors and a C-terminal mutant.
abstract: GluK1 is a kainate receptor subunit in the ionotropic glutamate receptor family and can form functional channels when expressed, for instance, in HEK-293 cells. However, the channel-opening mechanism of GluK1 is poorly understood. One major challenge to studying the GluK1 channel is its apparent low level of surface expression, which results in a low whole-cell current response even to a saturating concentration of agonist. A low level of surface expression is thought to be contributed by an endoplasmic reticulum (ER) retention signal sequence. When this sequence motif is present as in the C-terminus of wild-type GluK1-2b, the receptor is significantly retained in the ER. Conversely, when this sequence is either lacking, as in wild-type GluK1-2a (i.e., a different alternatively spliced isoform at the C-terminus), or disrupted, as in a GluK1-2b mutant (i.e., R896A, R897A, R900A, and K901A), there is a higher level of surface expression and a greater whole-cell current response. Here we characterize the channel-opening kinetic mechanism for these three GluK1 receptors expressed in HEK-293 cells by using a laser-pulse photolysis technique. Our results show that wild-type GluK1-2a, wild-type GluK1-2b, and the GluK1-2b mutant have identical channel opening and channel closing rate constants. These results indicate that the amino acid sequence near or within the C-terminal ER retention signal sequence, which affects receptor trafficking and/or expression, does not affect channel gating properties. Furthermore, as compared with the GluK2 kainate receptor, the GluK1 channel is faster to open, close, and desensitize by at least 2-fold, yet the EC(50) value of GluK1 is similar to that of GluK2.
authors.affiliation: Department of Chemistry and Center for Neuroscience Research, University at Albany, SUNY, Albany, New York 12222, United States.
authors.forename: Yan
authors.initials: Y
authors.lastname: Han
authors.forename: Congzhou
authors.initials: C
authors.lastname: Wang
authors.forename: Jae Seon
authors.initials: JS
authors.lastname: Park
authors.forename: Li
authors.initials: L
authors.lastname: Niu
mesh_terms: D017398:Alternative Splicing / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D018698:Glutamic Acid / Q000737:chemistry / Q000378:metabolism / Q000502:physiology; D057809:HEK293 Cells; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D009154:Mutation; D010446:Peptide Fragments / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D010782:Photolysis; D021381:Protein Transport / Q000235:genetics; D018092:Kainic Acid Receptors / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D015854:Up-Regulation / Q000235:genetics
chemical_list: D018698:Glutamic Acid; D007527:Isoenzymes; D010446:Peptide Fragments; D018092:Kainic Acid Receptors