From bedside‐to‐bench: What disease‐associated variants are teaching us about the NMDA receptor

Amin, Johansen; Moody, Gabrielle; Wollmuth, Lonnie P.

doi:10.1113/jp278705

Cited by 43 publications

(36 citation statements)

References 137 publications

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“…Topologically, the majority (about 80%) of GRIN1 , GRIN2A , and GRIN2B disease‐associated variants are located within TMD and LBD (see Figure 2 and Table S2). This result is in accordance with previous reports compiling the data from around 100 GRIN variants (Amin et al, 2020; XiangWei et al, 2018). This differential domain sensitivity is exhibited by all GluN subunits, in line with the high phylogenetic conservation degree and structural constraints of the NMDAR channel pore.…”

Section: Resultssupporting

confidence: 94%

“…By integrating the fragmented information collected from databases and research articles, the variant can finally be annotated as loss-of-function (LoF) and the corresponding clinical phenotype is displayed Topologically, the majority (about 80%) of GRIN1, GRIN2A, and GRIN2B disease-associated variants are located within TMD and LBD (see Figure 2 and Table S2). This result is in accordance with previous reports compiling the data from around 100 GRIN variants (Amin et al, 2020;XiangWei et al, 2018 (Esmenjaud et al, 2019). Conversely, the ATD presents a low association with neurological conditions, with occasional disease-associated variants located close to positive and negative allosteric binding sites.…”

Section: Grin Variants and Disease Associationsupporting

confidence: 93%

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GRIN database: A unified and manually curated repertoire of GRIN variants

et al. 2020

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Glutamatergic neurotransmission is crucial for brain development, wiring neuronal function, and synaptic plasticity mechanisms. Recent genetic studies showed the existence of autosomal dominant de novo GRIN gene variants associated with GRINrelated disorders (GRDs), a rare pediatric neurological disorder caused by N-methyl-D-aspartate receptor (NMDAR) dysfunction. Notwithstanding, GRIN variants identification is exponentially growing and their clinical, genetic, and functional annotations remain highly fragmented, representing a bottleneck in GRD patient's stratification. To shorten the gap between GRIN variant identification and patient stratification, we present the GRIN database (GRINdb), a publicly available, nonredundant, updated, and curated database gathering all available genetic, functional, and clinical data from more than 4000 GRIN variants. The manually curated GRINdb outputs on a web server, allowing query and retrieval of reported GRIN variants, and thus representing a fast and reliable bioinformatics resource for molecular clinical advice. Furthermore, the comprehensive mapping of GRIN variants' genetic and clinical information along NMDAR structure revealed important differences in GRIN variants' pathogenicity and clinical phenotypes, shedding light on GRIN-specific fingerprints. Overall, the GRINdb and web server is a resource for molecular stratification of GRIN variants, delivering clinical and investigational insights into GRDs.

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Section: Resultssupporting

confidence: 94%

Section: Grin Variants and Disease Associationsupporting

confidence: 93%

GRIN database: A unified and manually curated repertoire of GRIN variants

et al. 2020

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“…Although the mechanism of NMDAR gating remains poorly understood, it has been hypothesized that the linker tethering the ABD to the TMD is responsible for facilitating communication between these two domains (Schorge et al, 2005;Sobolevsky et al, 2009;Talukder et al, 2010;Talukder and Wollmuth, 2011;Karakas and Furukawa, 2014;Lee et al, 2014;Gibb et al, 2018;Amin et al, 2020). Thus far, the flexible and dynamic nature of the ABD-TMD linker has prevented high resolution structure of this region, and the structure of the fully open NMDAR pore remains elusive (Tajima et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

“…In a population of >140,000 healthy individuals (available from gnomad.broadinstitute.org), the GluN2A and GluN2B pre-M1 helices are devoid of missense variants, suggesting strong selection against variation in this region. By contrast, the pre-M1 helices of these subunits harbor multiple de novo mutations in patients with epilepsy, intellectual disabilities, or developmental delays ( Swanger et al, 2016 ; Ogden et al, 2017 ; XiangWei et al, 2018 ; Amin et al, 2020 ). Within the GluN2A and GluN2B subunits, functional analysis of these disease-associated mutations revealed changes to NMDAR kinetics, further implicating the pre-M1 helix in the gating mechanism ( Ogden et al, 2017 ; Gibb et al, 2018 ).…”

Section: Introductionmentioning

confidence: 99%

NMDA receptor channel gating control by the pre-M1 helix

McDaniel

Ogden

Kell

et al. 2020

Journal of General Physiology

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The NMDA receptor (NMDAR) is an ionotropic glutamate receptor formed from the tetrameric assembly of GluN1 and GluN2 subunits. Within the flexible linker between the agonist binding domain (ABD) and the M1 helix of the pore-forming transmembrane helical bundle lies a two-turn, extracellular pre-M1 helix positioned parallel to the plasma membrane and in van der Waals contact with the M3 helix thought to constitute the channel gate. The pre-M1 helix is tethered to the bilobed ABD, where agonist-induced conformational changes initiate activation. Additionally, it is a locus for de novo mutations associated with neurological disorders, is near other disease-associated de novo sites within the transmembrane domain, and is a structural determinant of subunit-selective modulators. To investigate the role of the pre-M1 helix in channel gating, we performed scanning mutagenesis across the GluN2A pre-M1 helix and recorded whole-cell macroscopic and single channel currents from HEK293 cell-attached patches. We identified two residues at which mutations perturb channel open probability, the mean open time, and the glutamate deactivation time course. We identified a subunit-specific network of aromatic amino acids located in and around the GluN2A pre-M1 helix to be important for gating. Based on these results, we are able to hypothesize about the role of the pre-M1 helix in other NMDAR subunits based on sequence and structure homology. Our results emphasize the role of the pre-M1 helix in channel gating, implicate the surrounding amino acid environment in this mechanism, and suggest unique subunit-specific contributions of pre-M1 helices to GluN1 and GluN2 gating.

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“…Our experiments highlight the critical role the LBD-TMD linkers play in regulating NMDAR activity. Notable are the S2-M4/M4 segments which display numerous disease-associated variants (Yuan et al, 2014;Amin et al, 2018;Vyklicky et al, 2018;Amin et al, 2020). Further, the extracellular position of S2-M4 compared to the membrane-embedded position of M4 provides a strategic advantage for designing new drug therapies (Shi et al, 2019).…”

Section: Resultsmentioning

confidence: 99%

NMDA receptors require multiple pre-opening gating steps for efficient synaptic activity

Amin

Gochman

et al. 2020

Preprint

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NMDA receptors (NMDAR) are glutamate-gated ion channels that mediate the majority of fast excitatory synaptic transmission in the nervous system. A central feature of NMDAR physiology is the opening of the ion channel driven by presynaptically-released glutamate. Using glutamate applications to outside-out patches containing a single NMDAR in the continuous presence of the co-agonist glycine, we find that agonist-bound receptors transition to the open state via two conformations, an 'unconstrained pre-active' state that can rapidly transition to the open state and contributes to synaptic events, and a 'constrained pre-active' state that requires more energy and hence time to open and does not contribute to fast signaling. To define how agonist binding might drive these conformations, we decoupled the ligand-binding domains from specific transmembrane segments for the GluN1 and GluN2A subunits. Displacements of the central pore-forming M3 segments define the energy of fast channel opening. However, to enter the unconstrained conformation and contribute to fast signaling, a peripheral helix, the GluN2 pre-M1, must be displaced before the M3 segments move. This pre-M1 displacement is facilitated by the flexibility of another nearby peripheral element, the GluN1 and GluN2A S2-M4. We conclude that peripheral structural elements -pre-M1 and S2-M4 -work in concert to remove constraints and prime the channel for rapid opening, thus facilitating fast synaptic transmission. 214 words. McDaniel et al., 2020). The S2-M4 linker/M4 segment also regulates gating (Amin et al., 2017;Yelshanskaya et al., 2017;Shi et al., 2019). Still, the energetics and timing of these non-pore forming elements to receptor gating remains incomplete.Here, we address the gating mechanism in NMDARs. At most synapses, NMDARs are obligate hetero-tetramers composed of two GluN1 and two GluN2 (A-D) subunits. To define conformational changes between agonist binding and pore opening, we rapidly applied glutamate to outside-out patches containing single NMDARs and assessed the time between when glutamate was first applied to when the channel first opened ('latency to 1 st opening', see Materials and Methods) (Aldrich et al., 1983). With this approach, we identified that wild type GluN1/GluN2A NMDARs, in the transition from agonist-bound closed to the open state, transitions through one of two pre-active states: an 'unconstrained' state that can rapidly transition to the open state and contributes to fast synaptic signaling, and a 'constrained' state that slowly transitions to the open state, presumably due to higher energy requirements, and does not contribute to fast signaling. Further, by decoupling the LBD from the TMD for each specific LBD-TMD linker (S1-M1, M3-S2, and S2-M4, Figure 1B & 1C) (Niu et al., 2004;Kazi et al., 2014), we show that the non-pore forming linkers uniquely influence pore-opening by altering the stability of these pre-active conformations. Thus, our results highlight the temporal and coordinated movements from agonist binding to io...

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From bedside‐to‐bench: What disease‐associated variants are teaching us about the NMDA receptor

Abstract: NMDA receptors (NMDARs) are glutamate-gated ion channels that contribute to nearly all brain processes. Not surprisingly then, genetic variations in the genes encoding NMDAR

Cited by 43 publications

References 137 publications

GRIN database: A unified and manually curated repertoire of GRIN variants

GRIN database: A unified and manually curated repertoire of GRIN variants

NMDA receptor channel gating control by the pre-M1 helix

NMDA receptors require multiple pre-opening gating steps for efficient synaptic activity

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