Neto proteins: exploring the structural basis of kainate receptor modulation

Kainate receptors (KARs) are ionotropic glutamate receptors (iGluRs) that presynaptically modulate neurotransmitter release at both excitatory and inhibitory synapses in addition to modulating neuronal excitability. Additionally, KARs are implicated in different pathological states, such as excitotoxicity and stroke. KARs co-assemble with Neuropilin and tolloid-like auxiliary subunits (Neto1 and Neto2), which modulate both KAR gating mechanisms and neuronal localization. The effect of Neto protein co-assembly with KARs is subunit (GluK1-5), as well as Neto isoform, specific. While the importance of auxiliary subunits in native iGluR function has been demonstrated, the structural basis of iGluR modulation by auxiliary subunits remains elusive. The objective of our study is to identify the structural determinants and cellular mechanisms that underlie Neto1 and Neto2 function as KAR auxiliary subunits. Understanding the structural determinants of Neto protein function would provide key insight into how KAR gating and neuronal localization mechanisms are modulated, in addition to providing potential targets for therapeutic intervention. Neto auxiliary subunits have a defined, modular structure and share sequence homology, yet have differential functionalities, making them a good model to study iGluR modulation by auxiliary subunits. Neto1 and Neto2 are comprised of two large extracellular CUB domains, followed by an LDLa module, a transmembrane region and an intracellular carboxyl-terminus. The presence of these well-delineated domains affords us opportunity to investigate specific, defined motifs and residues responsible for their unique effects, which will provide detailed insight into the subtleties of KAR function. In Specific Aim 1, we will identify the structural determinants of Neto1 and Neto2 that underlie their differential modulation of GluK1 receptor gating. In specific Aim 2, we will identify the structural determinants and cellular mechanism required for GluK1 receptor synaptic targeting by Neto2. This information will identify potential sites to allosterically modulate KAR function, which could be important in guiding the structure-driven and rational development of new drugs for the treatment of diseases such as stroke.