Synaptic adhesion protein ELFN1 is a selective allosteric modulator of group III metabotropic glutamate receptors in trans

Abstract

Functional characterization of the GPCR interactome has been focused predominantly on intracellular interactions, yet GPCRs are increasingly found in complex with extracellular proteins. Extracellular leucine-rich repeat fibronectin type III domain containing 1 (ELFN1) was recently reported to physically anchor mGluR6 and mGluR7 across retinal and hippocampal synapses, respectively; however, the consequence of transsynaptic interactions on properties and pharmacology of these receptors are unknown. In the current study, we explore the effects of ELFN1 on mGluR signaling and pharmacology. First, we established the binding specificity of ELFN1 and found it to be recruited selectively to all group III mGluRs (mGluR4, mGluR6, mGluR7, and mGluR8), but not other mGluR species. Using site-directed mutagenesis we mapped binding determinants of this interaction to two distinct sites on the ELFN1 ectodomain. To evaluate functional aspects of the interaction, we developed a transcellular signaling assay in reconstituted HEK293 cells which monitors changes in mGluR activity in one cell following its exposure to separate ELFN1-containing cells. Using this platform, we found that ELFN1 acts as an allosteric modulator of class III mGluR activity in suppressing cAMP accumulation: altering both agonist-induced and constitutive receptor activity. Using bioluminescence resonance energy transfer-based real-time kinetic assays, we established that ELFN1 alters the ability of mGluRs to activate G proteins. Our findings demonstrate that core properties of class III mGluRs can be altered via extracellular interactions with ELFN1 which serves as a transsynaptic allosteric modulator for these receptors. Furthermore, our unique assay platform opens avenues for exploring transcellular/transsynaptic pharmacology of other GPCR transcomplexes.

Elfn1-Induced Constitutive Activation of mGluR7 Determines Frequency-Dependent Recruitment of Somatostatin Interneurons

“…Elfn1-mGluR7 interactions generate a structurally induced constitutive GPCR activation, tonically suppressing pyr ¡ SOM transmission. A previous report indicates that Elfn1 induces negative allosteric modulation of G-protein signaling in response to glutamate (Dunn et al, 2018). Although Elfn1induced negative allosteric modulation is not explicitly tested here, a reduction in L-AP4 mediated synaptic suppression for WT versus Elfn1 KO provides support for these in vitro findings.…”

“…3K ), suggesting a divergence from the diffusible second messenger signaling cascades typically evoked by agonist activation. Indeed, a recent study demonstrates that Elfn1 interactions produce negative allosteric modulation of Type III mGluRs by suppressing G-protein coupling efficiency for G␣ i and G␣ o (Dunn et al, 2018). This series of observations is instead consistent with the fast, membrane delimited, voltage-sensitive suppression of calcium channels by G-protein coupled receptors (GPCRs) described by Hille and colleagues (Beech et al, 1992).…”

“…2C). Elfn1 coimmunoprecipitates mGluR7, as well as other Type III mGluRs, indicating that these proteins physically interact (Tomioka et al, 2014;Dunn et al, 2018). We assessed the effects of mGluR7 on synaptic transmission at pyr ¡ SOM synapses by antagonizing mGluR7 with the mGluR (Type III)-selective antagonist MSOP (100 M).…”

Transcriptomic Organization of Human Posttraumatic Stress Disorder

“…2B & C); it is also decreased in female dlPFC and dACC and is a key driver in a female-specific network ( Table S3). ELFN1 encodes a synaptic adhesion protein that acts as an allosteric modulator of group III metabotropic glutamate receptors 41 ; it is also required for synapse formation on SST GABA interneurons by recruiting presynaptic mGluR7 (metabotropic glutamate receptor 7) and GluK2 (glutamate receptor kainate 2) 42 . Notably, there are multiple GABA related transcripts that are significantly down-regulated key-drivers in the same network as ELFN1, including SST, PNOC (Prepronociceptin, a GABA interneuron maker), GAD2…”

Expression and distribution of Trophoblast Glycoprotein in the mouse retina

“…The N-terminal extracellular region of TPBG contains a heavily glycosylated leucine-rich repeat domain, a common site of protein-protein interactions (Zhao et al, 2014) and several similarly structured transmembrane proteins have recently been identified as vital for the development of the rod-RBC synapse or for localization of RBC synaptic transduction components. ELFN1, a synaptic adhesion LRR protein expressed in rod photoreceptors, forms trans-synaptic complexes with mGluR6 in RBC dendrites, and is required for the development of a functional synapse (Yan Cao et al, 2015;Dunn, Patil, Cao, Orlandi, & Martemyanov, 2018).…”