GluN1 Subunit in the Cellular Nucleus: History
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Contributor:
Terry A. McNearney
,
Karin N. Westlund

The N-methyl-D-aspartate (NMDA) glutamate receptors function as plasma membrane ionic channels and take part in very tightly controlled cellular processes activating neurogenic and inflammatory pathways. In particular, the NR1 subunit (new terminology: GluN1) is required for many neuronal and non-neuronal cell functions, including plasticity, survival, and differentiation. Protein tyrosine kinase inhibitors can effectively reduce (i) pain-related behavior, (ii) GluN1 subunit expression increases in the spinal cord, and (iii) the shift of GluN1 subunit from a cell membrane to nuclear localization. 

  • pain
  • nuclear translocation
  • nucleus
  • nucleolus
  • epigenetics
  • membrane trafficking

1. Introduction

The N-methyl-D-aspartate (NMDA) glutamate receptors function as plasma membrane ionic channels and are part of very tightly controlled cellular processes activating neurogenic and inflammatory pathways. A large body of studies supports their functions in both normal physiology and disease states with a focus on the NMDA NR1 subunit (new terminology GluN1 subunit [1]) function in psychiatric and neurologic conditions. NMDA receptors are activated by excitatory amino acid (EAA) agonists, glutamate, aspartate, and NMDA. NMDA receptors in low levels are essential for neuronal development, differentiation, learning, survival, and plasticity [2][3]. NMDA receptors play a critical role in pre- and post-synaptic plasticity, especially learning and memory [4]. However, when glutamate receptor agonists are present in excess, binding to NMDA receptors produces neuronal/CNS/PNS excitotoxicity, pathology, conditions of acute pain [5][6], and ongoing severe, intractable pain [7][8]. Conditional deletion of the GluN1 subunit in the spinal cord dorsal horn reduces injury-induced pain [9]. Beyond this is involvement in anxiety/depression [10], disease states (schizophrenia, Parkinson’s [11][12]), dementia [13][14][15], and seizures [16][17].
Earlier studies established that the GluN1 subunit anchor component was necessary for heterodimer formation, cellular trafficking, and nuclear localization that was functionally specific [18][19]. Four subunits assemble the glutamate receptors, and each heterodimer strictly requires the GluN1 subunit to anchor GluN2 or GluN3 subunits, comprising a functional ionic channel. GluN1 subunits bind combinations of GluN2A, 2B, and GluN3A and 3B subunits that form heterodimers to anchor these components on the cellular plasma membrane [20][21][22]. The binding composition is noted as two obligatory GluN1 subunits, of which there are now reported at least eight distinct variant subunits and four or two variable subunits from the GluN2 (GluN2A-2D) and GluN3 (GluN3A-3B), respectively, producing 8 × 6 = 48 potential heterodimers.
The GluN1-1a subunits have a nuclear localization sequence (NLS) exclusive of tissue specificity [18][23]. Additional studies have reported the alternative splicing of messenger RNA producing similar proteins that target different tissues, determine its functional fate [24], and have expanded studies to include expression and functioning in non-neuronal tissues. Much of the recent research has focused on characterizing the GluN2 subunits of the GluN1/GluN2 heterodimer. Shifts in the composition of the heterodimer, based on GluN2 or N3 subunits, denote the functional roles of the receptor complex and promote the neuroplasticity of the glutaminergic system throughout the CNS [23][24].

2. Potential Fates of GluN1 Subunit in the Cellular Nucleus

2.1. GluN1 Subunit Signaling Induces Nuclear Translocation

GluN1 subunit occupation in the cell nucleus has been reported since early 2000, and a resurgence of research is providing a better understanding to fill in the gaps. This has included reports of the bipartite NLS contained in the GluN1-1a subunit [18][25], protein isoforms, and sequence cassettes. For example, the GluN1/N2A/B functional tetramer is inserted into the cellular plasma membrane site as an ionic channel. When activated, the heterodimer is phosphorylated at the GluN2 cytoplasmic regions, and the GluN1 subunit transmembrane region protein undergoes intermembrane proteolysis, cleaving the cytoplasmic portion in the cell membrane. The heterodimer internalizes and translocates from the cell’s plasma membrane to the cell’s nuclear membrane [18][26]. This region contains a bipartite NLS with two clusters of short sequences of basic amino acids, mainly lysines (K) and arginines (R), separated by a link of a variable number of amino acids. The two regions of basic amino acids on the protein surface comprise the regions recognized for binding along the nuclear membrane and passing into the nucleus. In the case of GluN1-1a, the NLS sequence regions with basic amino acids in bold type are:
KRHK-spacer region—KKKATFRAITSTLASSFKRRR [25].

2.2. GluN1 Subunit Staining in the Cellular Nucleus

The nuclear translocation of the GluN1 subunit after glutamate activation suggests it also plays a direct role in the fast intracellular signaling responses to extracellular glutamate activation. Long-term potentiation (LTP) mediated facilitation requires an active importin nuclear import pathway [27]. Examination of the function of the GluN1 subunit in nuclear translocation has found the C1 domain binds calmodulin to assist calcium entry into the nucleus [19][23]. This regulates LTP and synaptic plasticity in hippocampal cultures. Glutamate-mediated (LTP-like) overactivation in pain states likely involves calcium entry into the nucleus through ion channels that include GluN1 subunits. The evidence for this appears below.
Detection of increased cell nuclear staining of GluN1 subunit in activated cells is easily appreciated in vivo or in vitro. For example, increased cell nuclear staining of the GluN1 subunit was observed with visual microscopic examination of human clonal neuroblastoma cell cultures (SH-SY5Y) activated with glutamate or NMDA for 4 h, and nuclear staining was inhibited by preincubation with active NMDA protein tyrosine kinase (PTK) inhibitors genistein or staurosporin in vitro [28]. Increased nuclear staining was also easily observed by 2 h after incubation with NMDA and ACPD in human synovial fibroblast cells [29]. Additionally, staining of the GluN1 subunit on the nuclear rim and nucleoplasm was appreciated in the presence of cycloheximide [28], demonstrating that nuclear localization is an incitement event for the subunit and not solely the purview of newly synthesized GluN1 subunit.
In complementary studies inducing synovial joint capsule inflammation, secondary tactile allodynia in the same hindlimb resulted in the same time span of two hours following excess glutamate released into the joint capsule from activated peripheral nerves in a rat intra-articular K/C arthritis model [30]. The afferent nerve and spinal glutamate neurons release glutamate into the spinal cord dorsal horn (SCDH) in response to activation or injury [31]. Models providing information on nociception/pain study either acute pain, where glutamate receptor stimulation is limited timewise, or chronic pain, where the glutamate receptor is overstimulated long-term. Many molecular, epigenetic, and inflammatory events are ongoing, accompanied by prolonged exposure to glutamate and overactivation of glutamate receptors. Continued activation of the glutamate receptors results in excessive intracellular chloride that increases the neuronal membrane potential above the threshold, resulting in the reversal of GABA from inhibitory to excitatory continuous firing, contributing to chronic pain [32][33].

3. Roles and Potential Fates of GluN1 Subunit in The Cellular Nucleus

The regional and functional assignments provided in the GluN1/2 subunit heterodimers have been attributed mostly to the GluN2 subunits, after they are trafficked and delivered to the nucleus via the GluN1 receptor subunit. However, the GluN1 subunit has several indicators of its own potential to influence heterodimer and/or subnuclear organelle assignments.
1. The putative NLS and/or nucleolar localization sequences (NoLS) located in cassette 1 of the GluN1 subunit has two short regions rich in basic amino acids, which comprise the NLS and are potentially involved in nucleolar signaling [34][35]. GluN1 subunits are tightly subjected to multiple levels of regulation, affecting subunit expression, subcellular location, and assembly of functional receptors, and their signaling complexes [36].
2. The gene for the GluN1 subunit is expressed in early development in virtually all neurons, and is transcriptionally upregulated during neuronal differentiation. NMDA agonists and EAAs increase cellular GluN1 subunit levels, as demonstrated with widespread cell membrane, intracellular, and nuclear GluN1 subunit staining. GluN1 subunit nuclear translocalization is reported for both human synoviocytes and for rat spinal cord nociceptive neurons at light and EM levels [28][37]. Nuclear translocation of GluN1 is reported for neurons in eye tissue [38][39]. Additionally, GluN1 subunit activation has been reported in models of ischemia, neurogenic, and inflammatory responses, which also stimulate EAA release and increase GluN1 subunit expression. Zhou and Duan have reported that both GluN1 and GluN2 are needed to translocate [23].
3. There is close coordination in neurons between the assembly of functional heteromeric to tetrameric receptors and the fates of these individual subunits. In addition, two pools of mRNA for the GluN1 subunit have been reported with distinct translational activities. These generate two stores of GluN1 subunits that are differentially assembled with GluN2 subunits to form heterodimers with distinct functions and turnover rates, providing an additional and possibly tissue-specific level of control for protein turnover and trafficking [20]. Nuclear membrane translocation reportedly occurs by endocytotic and de novo mechanisms. Activity dependent clathrin mediated internalization of GluN1 subunit is reported [26]. Nociceptive neurons are overactivated in pain states with increased GluN1 subunit cellular and nuclear ring immunostaining [28]. Staining is greatly reduced by tyrosine kinase inhibition. Post-transcriptional mechanisms also contribute to GluN1 subunit regulation in brain development [40][41]. Studies have also reported the importance of post-translational histone modifications in epigenetic transcriptional control of nociceptive pathways [42][43].
4. Structurally, the GluN1 subunit promoter region is located directly upstream of the transcriptional start site (TSS) and is exceptional for the number of transcriptionally reactive binding regions close to the 3’ site. Transcriptionally active binding regions have been reported for SP-1, NFkB, MEF-2, GC-rich regions, CREB, REI, AP-1, egr-1, and ARC binding regions [5]. The GluN1 promoter allows a spectrum of potential responses, such as activation, de-repression, or suppression of downstream transcribable sequences.
5. GluN1 subunit activation is reported to consequently activate Immediate Early Genes (IEG), e.g., c-fos, zif268, and egr-1 [44][45], and conversely, IEG products are reported to activate GluN subunits. The term IEG describes viral regulatory proteins or cellular proteins generated immediately following stimulation of a resting cell by internal or external stimuli, triggering immediate gene transcription that does not require de novo protein synthesis. IEG products are usually transcription factors, which are DNA-binding protein activators of signaling pathways. They are rapidly and transiently activated to respond to a plethora of cellular and extracellular stimuli, serving as an important cellular first response system. The GluN1 subunit can direct downstream nuclear functioning via nuclear DNA binding sites, immediate early gene products, cytoplasmic input, and environmental signals.

This entry is adapted from the peer-reviewed paper 10.3390/ijms241713196

References

  1. Harding, S.D.; Armstrong, J.F.; Faccenda, E.; Southan, C.; Alexander, S.P.H.; Davenport, A.P.; Pawson, A.J.; Spedding, M.; Davies, J.A.; Iuphar, N. The IUPHAR/BPS guide to PHARMACOLOGY in 2022: Curating pharmacology for COVID-19, malaria and antibacterials. Nucleic Acids Res. 2021, 50, D1282–D1294.
  2. Marini, A.M.; Rabin, S.J.; Lipsky, R.H.; Mocchetti, I. Activity-dependent Release of Brain-derived Neurotrophic Factor Underlies the Neuroprotective Effect of N-Methyl-d-aspartate. J. Biol. Chem. 1998, 273, 29394–29399.
  3. Hansen, K.B.; Wollmuth, L.P.; Bowie, D.; Furukawa, H.; Menniti, F.S.; Sobolevsky, A.I.; Swanson, G.T.; Swanger, S.A.; Greger, I.H.; Nakagawa, T.; et al. Structure, Function, and Pharmacology of Glutamate Receptor Ion Channels. Pharmacol. Rev. 2021, 73, 1469–1658.
  4. Chandrasekar, R. Alcohol and NMDA receptor: Current research and future direction. Front. Mol. Neurosci. 2013, 6, 14.
  5. Bai, G.; Hoffman, P.W. Chapter 5 Biology of the NMDA Receptor; Van Dongen, A.M., Ed.; CRC Press; Taylor & Francis: Boca Raton, FL, USA, 2009.
  6. Willis, W.D., Jr. The role of TRPV1 receptors in pain evoked by noxious thermal and chemical stimuli. Exp. Brain Res. 2009, 196, 5–11.
  7. Kristensen, J.D.; Svensson, B.; Gordh, T. The NMDA-receptor antagonist CPP abolishes neurogenic ‘wind-up pain’ after intrathecal administration in humans. Pain 1992, 51, 249–253.
  8. Riccardi, A.; Guarino, M.; Serra, S.; Spampinato, M.D.; Vanni, S.; Shiffer, D.; Voza, A.; Fabbri, A.; De Iaco, F.; on behalf of the Study and Research Center of the Italian Society of Emergency Medicine. Narrative Review: Low-Dose Ketamine for Pain Management. J. Clin. Med. 2023, 12, 3256.
  9. South, S.M.; Kohno, T.; Kaspar, B.K.; Hegarty, D.; Vissel, B.; Drake, C.T.; Ohata, M.; Jenab, S.; Sailer, A.W.; Malkmus, S.; et al. A Conditional Deletion of the NR1 Subunit of the NMDA Receptor in Adult Spinal Cord Dorsal Horn Reduces NMDA Currents and Injury-Induced Pain. J. Neurosci. 2003, 23, 5031–5040.
  10. Hietamies, T.M.; McInnes, L.A.; Klise, A.J.; Worley, M.J.; Qian, J.J.; Williams, L.M.; Heifets, B.D.; Levine, S.P. The effects of ketamine on symptoms of depression and anxiety in real-world care settings: A retrospective controlled analysis. J. Affect. Disord. 2023, 335, 484–492.
  11. Yang, J.; Hertz, E.; Zhang, X.; Leinartaité, L.; Lundius, E.G.; Li, J.; Svenningsson, P. Overexpression of α-synuclein simultaneously increases glutamate NMDA receptor phosphorylation and reduces glucocerebrosidase activity. Neurosci. Lett. 2016, 611, 51–58.
  12. Zhang, Z.; Zhang, S.; Fu, P.; Zhang, Z.; Lin, K.; Ko, J.K.-S.; Yung, K.K.-L. Roles of Glutamate Receptors in Parkinson’s Disease. Int. J. Mol. Sci. 2019, 20, 4391.
  13. Lewerenz, J.; Maher, P. Chronic Glutamate Toxicity in Neurodegenerative Diseases—What Is the Evidence? Front. Neurosci. 2015, 9, 469.
  14. Lipton, S.A. NMDA Receptors, Glial Cells, and Clinical Medicine. Neuron 2006, 50, 9–11.
  15. Nagu, P.; Parashar, A.; Behl, T.; Mehta, V. Gut Microbiota Composition and Epigenetic Molecular Changes Connected to the Pathogenesis of Alzheimer’s Disease. J. Mol. Neurosci. 2021, 71, 1436–1455.
  16. Rodríguez-Muñoz, M.; Sánchez-Blázquez, P.; Garzón, J. Fenfluramine diminishes NMDA receptor-mediated seizures via its mixed activity at serotonin 5HT2A and type 1 sigma receptors. Oncotarget 2018, 9, 23373–23389.
  17. Levite, M.; Goldberg, H. Autoimmune Epilepsy—Novel Multidisciplinary Analysis, Discoveries and Insights. Front. Immunol. 2022, 12, 762743.
  18. Holmes, K.D.; Mattar, P.; Marsh, D.R.; Jordan, V.; Weaver, L.C.; Dekaban, G.A. The C-terminal C1 cassette of the N-methyl-d-aspartate receptor 1 subunit contains a bi-partite nuclear localization sequence. J. Neurochem. 2002, 81, 1152–1165.
  19. Zhou, L.; Duan, J. The NMDAR GluN1-1a C-terminus binds to CaM and regulates synaptic function. Biochem. Biophys. Res. Commun. 2021, 534, 323–329.
  20. Ulbrich, M.H.; Isacoff, E.Y. Rules of engagement for NMDA receptor subunits. Proc. Natl. Acad. Sci. USA 2008, 105, 14163–14168.
  21. Jeffrey, R.A.; Ch’Ng, T.H.; O’Dell, T.J.; Martin, K.C. Activity-Dependent Anchoring of Importin α at the Synapse Involves Regulated Binding to the Cytoplasmic Tail of the NR1-1a Subunit of the NMDA Receptor. J. Neurosci. 2009, 29, 15613–15620.
  22. Niu, M.; Yang, X.; Li, Y.; Sun, Y.; Wang, L.; Ha, J.; Xie, Y.; Gao, Z.; Tian, C.; Wang, L.; et al. Progresses in GluN2A-containing NMDA Receptors and their Selective Regulators. Cell. Mol. Neurobiol. 2023, 43, 139–153.
  23. Zhou, L.; Duan, J. The C-terminus of NMDAR GluN1-1a Subunit Translocates to Nucleus and Regulates Synaptic Function. Front. Cell. Neurosci. 2018, 12, 334.
  24. Garraway, S.M.; Xu, Q.; Inturrisi, C.E. Design and Evaluation of Small Interfering RNAs That Target Expression of the N-Methyl-d-aspartate Receptor NR1 Subunit Gene in the Spinal Cord Dorsal Horn. Experiment 2007, 322, 982–988.
  25. Holmes, K.D.; Mattar, P.A.; Marsh, D.R.; Weaver, L.C.; Dekaban, G.A. The N-Methyl-d-aspartate Receptor Splice Variant NR1–4 C-terminal Domain. Deletion analysis and role in subcellular distribution. J. Biol. Chem. 2002, 277, 1457–1468.
  26. Cheng, F.; Li, X.; Li, Y.; Wang, C.; Wang, T.; Liu, G.; Baskys, A.; Uéda, K.; Chan, P.; Yu, S. α-Synuclein promotes clathrin-mediated NMDA receptor endocytosis and attenuates NMDA-induced dopaminergic cell death. J. Neurochem. 2011, 119, 815–825.
  27. Thompson, K.R.; Otis, K.O.; Chen, D.Y.; Zhao, Y.; O’Dell, T.J.; Martin, K.C. Synapse to Nucleus Signaling during Long-Term Synaptic Plasticity: A Role for the Classical Active Nuclear Import Pathway. Neuron 2004, 44, 997–1009.
  28. Westlund, K.N.; Lu, Y.; Zhang, L.; Pappas, T.C.; Zhang, W.-R.; Taglialatela, G.; McIlwrath, S.L.; McNearney, T.A. Tyrosine Kinase Inhibitors Reduce NMDA NR1 Subunit Expression, Nuclear Translocation, and Behavioral Pain Measures in Experimental Arthritis. Front. Physiol. 2020, 11, 440.
  29. McNearney, T.A.; Ma, Y.; Chen, Y.; Taglialatela, G.; Yin, H.; Zhang, W.-R.; Westlund, K.N. A peripheral neuroimmune link: Glutamate agonists upregulate NMDA NR1 receptor mRNA and protein, vimentin, TNF-α, and RANTES in cultured human synoviocytes. Am. J. Physiol. Integr. Comp. Physiol. 2010, 298, R584–R598.
  30. Lawand, N.B.; McNearney, T.; Westlund, K.N. Amino acid release into the knee joint: Key role in nociception and inflammation. Pain 2000, 86, 69–74.
  31. Sorkin, L.; Westlund, K.; Sluka, K.; Dougherty, P.; Willis, W. Neural changes in acute arthritis in monkeys. IV. Time-course of amino acid release into the lumbar dorsal horn. Brain Res. Rev. 1992, 17, 39–50.
  32. Pol, A.N.v.D.; Obrietan, K.; Chen, G. Excitatory Actions of GABA after Neuronal Trauma. J. Neurosci. 1996, 16, 4283–4292.
  33. Coull, J.A.M.; Boudreau, D.; Bachand, K.; Prescott, S.A.; Nault, F.; Sík, A.; De Koninck, P.; De Koninck, Y. Trans-synaptic shift in anion gradient in spinal lamina I neurons as a mechanism of neuropathic pain. Nature 2003, 424, 938–942.
  34. Kundu-Michalik, S.; Bisotti, M.-A.; Lipsius, E.; Bauche, A.; Kruppa, A.; Klokow, T.; Kammler, G.; Kruppa, J. Nucleolar Binding Sequences of the Ribosomal Protein S6e Family Reside in Evolutionary Highly Conserved Peptide Clusters. Mol. Biol. Evol. 2008, 25, 580–590.
  35. Kass-Simon, G.; Zompa, M.A.; Scappaticci, A.A.; Zackroff, R.V.; Hufnagel, L.A. Nucleolar binding of an anti-NMDA receptor antibody in hydra: A non-canonical role for an NMDA receptor protein? J. Exp. Zool. Part A Ecol. Integr. Physiol. 2009, 311, 763–775.
  36. Gascón, S.; Deogracias, R.; Sobrado, M.; Roda, J.M.; Renart, J.; Rodríguez-Peña, A.; Díaz-Guerra, M. Transcription of the NR1 Subunit of the N-Methyl-d-aspartate Receptor Is Down-regulated by Excitotoxic Stimulation and Cerebral Ischemia. J. Biol. Chem. 2005, 280, 35018–35027.
  37. Ye, Z.; Westlund, K.N. Ultrastructural localization of glutamate receptor subunits (NMDAR1, AMPA GluR1 and GluR2/3) and spinothalamic tract cells. Neuroreport 1996, 7, 2581–2586.
  38. Fan, W.; Xing, Y.; Zhong, Y.; Chen, C.; Shen, Y. Expression of NMDA receptor subunit 1 in the rat retina. Acta Histochem. 2013, 115, 42–47.
  39. Ishimaru, Y.; Sumino, A.; Kajioka, D.; Shibagaki, F.; Yamamuro, A.; Yoshioka, Y.; Maeda, S. Apelin protects against NMDA-induced retinal neuronal death via an APJ receptor by activating Akt and ERK1/2, and suppressing TNF-α expression in mice. J. Pharmacol. Sci. 2017, 133, 34–41.
  40. Del Caño, G.G.; Gerrikagoitia, I.; Sarasa, M.; Matute, C.; Martínez-Millán, L. Ionotropic glutamate receptor subunits are differentially regulated in the motoneuronal pools of the rat hypoglossal nucleus in response to axotomy. J. Neurocytol. 2000, 29, 509–523.
  41. Awobuluyi, M.; Lipton, S.A.; Sucher, N.J. Translationally distinct populations of NMDA receptor subunit NR1 mRNA in the developing rat brain. J. Neurochem. 2003, 87, 1066–1075.
  42. Ghosh, K.; Pan, H.-L. Epigenetic Mechanisms of Neural Plasticity in Chronic Neuropathic Pain. ACS Chem. Neurosci. 2022, 13, 432–441.
  43. Irfan, J.; Febrianto, M.R.; Sharma, A.; Rose, T.; Mahmudzade, Y.; Di Giovanni, S.; Nagy, I.; Torres-Perez, J.V. DNA Methylation and Non-Coding RNAs during Tissue-Injury Associated Pain. Int. J. Mol. Sci. 2022, 23, 752.
  44. Loureiro, C.M.; Fachim, H.A.; Corsi-Zuelli, F.; Shuhama, R.; Joca, S.; Menezes, P.R.; Dalton, C.F.; Del-Ben, C.M.; Louzada-Junior, P.; Reynolds, G.P. Epigenetic-mediated N-methyl-D-aspartate receptor changes in the brain of isolated reared rats. Epigenomics 2020, 12, 1983–1997.
  45. Guglietti, B.; Sivasankar, S.; Mustafa, S.; Corrigan, F.; Collins-Praino, L.E. Fyn Kinase Activity and Its Role in Neurodegenerative Disease Pathology: A Potential Universal Target? Mol. Neurobiol. 2021, 58, 5986–6005.
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