GluA1 signal peptide determines the spatial assembly of heteromeric AMPA receptors

Abstract

AMPA-type glutamate receptors (AMPARs) mediate fast excitatory neurotransmission and predominantly assemble as heterotetramers in the brain. Recently, the crystal structures of homotetrameric GluA2 demonstrated that AMPARs are assembled with two pairs of conformationally distinct subunits, in a dimer of dimers formation. However, the structure of heteromeric AMPARs remains unclear. Guided by the GluA2 structure, we performed cysteine mutant cross-linking experiments in fulllength GluA1/A2, aiming to draw the heteromeric AMPAR architecture. We found that the amino-terminal domains determine the first level of heterodimer formation. When the dimers further assemble into tetramers, GluA1 and GluA2 subunits have preferred positions, possessing a 1-2-1-2 spatial assembly. By swapping the critical sequences, we surprisingly found that the spatial assembly pattern is controlled by the excisable signal peptides. Replacements with an unrelated GluK2 signal peptide demonstrated that GluA1 signal peptide plays a critical role in determining the spatial priority. Our study thus uncovers the spatial assembly of an important type of glutamate receptors in the brain and reveals a novel function of signal peptides.AMPA receptors | signal peptide | spatial assembly | stoichiometry

Signal peptide represses GluK1 surface and synaptic trafficking through binding to amino-terminal domain

“…However, growing evidence has revealed postcleavage physiological functions for signal peptides 21 , 22 . Our recent work has identified a role of the GluA1 signal peptide in determining the spatial assembly of heteromeric AMPA receptors 23 , which clearly functions in a cis manner. Here we uncover a function of the signal peptide of GluK1, which binds to the receptor like a ligand and is involved in the regulation of KAR intracellular trafficking in a trans manner.…”

“…Besides the well-characterized roles in ER targeting and membrane insertion 20 , 21 , some signal peptides have post-targeting functions, either as transmembrane peptides, or released into the cytosol or ER lumen after intramembrane proteolysis 22 . Recently, we have found that the signal peptide of AMPA receptor subunit GluA1 has an unconventional function of regulating the subunit spatial position for heteromeric GluA1/A2 receptor assembly 23 , suggesting that signal peptides of glutamate receptors might have other cellular and molecular functions in addition to their canonical ER targeting roles.…”

Architecture of the heteromeric GluA1/2 AMPA receptor in complex with the auxiliary subunit TARP γ8

“…GluA2 was detected in both AC and BD positions of the GluA2/3 receptor (25), raising the possibility that the subunit arrangement in GluA2/3 receptors could be more flexible. Placement rules, perhaps controlled by signal sequence parameters on initial assembly (33), will have consequences for gating control, modulation by auxiliary subunits and organization of the NTD layer (25), a platform for AMPAR positioning at synapses (68, 69). Lastly, we hypothesize that the asymmetric distribution of lipid-like densities, proximal to the γ8 β1 and β5 strands, is caused by TARP γ8 and may point to an additional regulatory capacity of auxiliary subunits, either by influencing the local lipid environment of synaptic receptors, or by modulation of channel properties through lipids, analogous to KARs (53).…”

Structural and Functional Architecture of AMPA-Type Glutamate Receptors and Their Auxiliary Proteins

“…Interestingly however, crosslinking experiments suggest that this positioning may not be strictly adhered to, as GluA2 can be located either proximal or distal to the pore axis in both GluA2/3 and GluA2/4 AMPARs (Herguedas et al, 2016). In apparent contrast, a recent study reported positional preference in GluA1/2 heteromers, where the GluA1 subunits are located pore-proximal, a feature driven by sequence elements in the signal peptide (He et al, 2016).…”