Channel-opening kinetic mechanism of wild-type GluK1 kainate receptors and a C-terminal mutant.

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.