Exploring the Molecular Mechanisms and Trafficking of Kainate Receptors

Student thesis: Doctoral ThesisDoctor of Philosophy (PhD)

Abstract

Kainate receptors (KARs) and AMPA receptors (AMPARs) are tetrameric glutamate-gated ion channels that play critical roles in neuronal excitability, signal transduction and synaptic plasticity. Their dysfunction has been strongly implicated in neurological, psychiatric, and neurodegenerative disorders including Alzheimer’s disease, autism, and epilepsy. However, the precise inter-relationship and relative synaptic localisations of KAR and AMPAR complexes, and crosstalk between their signalling, are not well understood. KAR activation can bidirectionally regulate AMPAR surface expression and synaptic strength. To explore the molecular pathways that underpin KAR regulation of AMPARs investigated whether the receptor complexes and/or individual subunits interact using co-immunoprecipitation techniques in HEK293T cells by exogenously expressing KAR and AMPAR subunits, and also in rat cortical neurons overexpressing KARs. AMPAR subunit GluA1 co-immunoprecipitated with the KAR subunit GluK2 in both endogenous and heterologous systems. Furthermore, contrary to previous reports, KARs and AMPARs localised to the same dendritic spines at the same time. Building on these initial data tested the hypothesis that, at least at some synapses, KARs regulate AMPARs either by binding of individual AMPAR and KAR subunits or, perhaps more likely, through common interacting proteins.
The expression of KAR on the surface is closely regulated, via GluK2 subunit post-translational modifications (PTMs). When GluK2-type KARs are activated by an agonist, GluK2 is phosphorylated at S868, which encourages SUMOylation at K886 and receptor endocytosis. Moreover, palmitoylation of GluK2 has been demonstrated. It is unclear, nevertheless, how SUMOylation, phosphorylation, and palmitoylation interact to coordinate KAR trafficking. Utilizing a collection of GluK2 mutants specific to specific sites, I examined the correlation among GluK2 PTMs and their influence on the expression of KAR surface. The basal palmitoylation of GluK2 is reduced upon stimulation with kainate (KA). Additionally, a non-palmitoylatable GluK2 mutant (C858/C871A) is resistant to KA-induced internalization and exhibits increased basal S868 phosphorylation and K886 SUMOylation. These findings suggest that palmitoylation of GluK2 plays a role in stabilizing KARs.
Date of Award7 May 2024
Original languageEnglish
Awarding Institution
  • The University of Bristol
SupervisorJeremy M Henley (Supervisor)

Keywords

  • kainate receptors
  • palmitoylation
  • Phosphorylation
  • ampa receptors
  • SUMOylation
  • Receptor trafficking

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