Differences in interactions between transmembrane domains tune the activation of metabotropic glutamate receptors

Abstract

The metabotropic glutamate receptors (mGluRs) form a family of neuromodulatory G protein-coupled receptors that contain both a seven-helix transmembrane domain (TMD) and a large extracellular ligand-binding domain (LBD) which enables stable dimerization. While numerous studies have revealed variability across subtypes in the initial activation steps at the level of LBD dimers, an understanding of inter-TMD interaction and rearrangement remains limited. Here we use a combination of single molecule fluorescence, molecular dynamics, functional assays, and conformational sensors to reveal that distinct TMD assembly properties drive differences between mGluR subtypes. We uncover a variable region within transmembrane helix 4 (TM4) that contributes to homo- and heterodimerization in a subtype-specific manner and tunes orthosteric, allosteric and basal activation. We also confirm a critical role for a conserved inter-TM6 interface in stabilizing the active state during orthosteric or allosteric activation. Together this study informs a working model of inter-TMD rearrangement that drives mGluR function.

Structural and compositional diversity in the kainate receptor family

“…Defining the stoichiometry and subunit arrangement of KAR di-heteromers Having established the general structural compatibility (Figure 1) and extensive co-expression of KAR subunit types (Figure 2), we aimed to assess the assembly, stoichiometry, and subunit arrangement of KARs composed of different pairs of subunit types, starting with di-heteromers. We employed the SiMPull method in which detergent-solubilized and fluorophore-tagged receptors are immobilized via antibodies and their subunits counted by photobleaching step analysis (Jain et al, 2011;Lee et al, 2020;Levitz et al, 2016;Royal et al, 2019;Thibado et al, 2021). We first generated GluK1 and GluK2 constructs with an N-terminal hemagglutinin (HA) epitope followed by a SNAP tag (Figure S4B) amenable to labeling with organic dyes, and both maintained normal receptor function (Figure S4B; Table S2).…”

Living on the edge

“…The assembly process of protomers into functional dimers remains poorly understood, but high-resolution mGluR structures and mutational analyses suggest that dimerization relies primarily on interactions between the hydrophobic interfaces of the VFT and interactions between the TMDs (El Moustaine et al, 2012;Koehl et al, 2019;Kunishima et al, 2000;Levitz et al, 2016). Interestingly, interactions between the TMDs play a predominant role in homodimerization of mGluR2 while for other mGluRs TMD interactions play only a moderate role in receptor dimerization (Gutzeit et al, 2019;Thibado et al, 2021). While mGluRs were long thought to be strict homodimeric receptors, a number of recent studies indicate that mGluRs are also able to form various heterodimer combinations (Doumazane et al, 2011;Habrian et al, 2019;Kammermeier, 2012;Levitz et al, 2016;Moreno Delgado et al, 2017;Pandya et al, 2016;Werthmann et al, 2020;Yin et al, 2014).…”