Stargazin Differentially Controls the Trafficking of -Amino-3- hydroxyl-5-methyl-4-isoxazolepropionate and Kainate Receptors

Synaptic plasticity at excitatory synapses in the brain is largely achieved by rapid changes in the number of synaptic -amino3-hydroxyl-5-methyl-4-isoxazolepropionate (AMPA) receptors. Stargazin, a membrane protein that interacts with AMPA receptors, is believed to play a pivotal role in trafficking AMPA receptors to the plasma membrane and targeting them to the synapse. However, it is unclear whether the trafficking of kainate receptors, which are structurally very similar to AMPA receptors, is also dependent on stargazin. Here we show that in both cerebellar granule cells and in Xenopus laevis oocytes expression system, surface delivery of kainate receptor is independent of stargazin. These results suggest that stargazin action is highly selective for AMPA receptors. Glutamate, the transmitter released from excitatory synapses in the brain, acts on three subtypes of ionotropic receptors: AMPA, NMDA, and kainate receptors. Synaptic AMPA receptors, in contrast to NMDA receptors, are highly mobile (Song et al., 1998; Luscher et al., 1999; Luthi et al., 1999; Noel et al., 1999; Ziff, 1999; Sheng, 2001; Malinow and Malenka, 2002), and changes in the number of synaptic AMPA receptors play an essential role in synaptic plasticity (Luscher et al., 1999; Daw et al., 2000; Malinow et al., 2000; Lu et al., 2001). We showed previously (Chen et al., 2000) that the delivery of AMPA receptors to the surface of cerebellar granule cells requires stargazin, the four-pass transmembrane protein defective in the ataxic stargazer mouse. On the other hand, the trafficking of NMDA receptors in these cells is independent of stargazin, highlighting the fundamental difference in the mechanisms involved in the delivery of these two subtypes of glutamate receptors to the membrane surface. In contrast to the abundant literature on the trafficking of AMPA and NMDA receptors, little is known about kainate receptor trafficking. In particular, a role for stargazin in this process has not been addressed. Because kainate receptors are structurally very similar to AMPA receptors, such information will provide important insight into the selectivity of the action of stargazin. To address this issue, we used the Xenopus laevis oocyte expression system, which allowed us to measure quantitatively the influence of stargazin on the surface expression of glutamate receptors. This system also allowed us to test whether stargazin effects on AMPA receptor trafficking can occur without other neuron-specific components. Materials and Methods Constructs. Stargazin, -1, and glutamate receptor subunits were subcloned into pGEM-HE vector (a gift from Lily Jan, University of California San Francisco). In vitro transcription of cRNAs was done using AmpliScribe T7 transcription kit (Epicentre Technolo-