Glutamate Acts as a Partial Inverse Agonist to Metabotropic Glutamate Receptor with a Single Amino Acid Mutation in the Transmembrane Domain

Yanagawa, Masataka; Yamashita, Takahiro; Shichida, Yoshinori

doi:10.1074/jbc.m112.437780

Cited by 13 publications

(14 citation statements)

References 36 publications

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“…We next measured the G protein activation ability of the visual pigments by a [ 35 S]GTPγS binding assay in complete darkness (under infrared light). The [ 35 S]GTPγS binding assay is a commonly used technique to evaluate the G protein activation rate of G protein-coupled receptors by measuring the GDP-GTPγS exchange rate of G proteins in vitro 11 (See Methods). The concentrations of visual pigments in the aliquots were compared by Western blotting analysis ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Origin of the low thermal isomerization rate of rhodopsin chromophore

Yanagawa

Kojima

Yamashita

et al. 2015

Sci Rep

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Low dark noise is a prerequisite for rod cells, which mediate our dim-light vision. The low dark noise is achieved by the extremely stable character of the rod visual pigment, rhodopsin, which evolved from less stable cone visual pigments. We have developed a biochemical method to quickly evaluate the thermal activation rate of visual pigments. Using an isomerization locked chromophore, we confirmed that thermal isomerization of the chromophore is the sole cause of thermal activation. Interestingly, we revealed an unexpected correlation between the thermal stability of the dark state and that of the active intermediate MetaII. Furthermore, we assessed key residues in rhodopsin and cone visual pigments by mutation analysis and identified two critical residues (E122 and I189) in the retinal binding pocket which account for the extremely low thermal activation rate of rhodopsin.

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Section: Resultsmentioning

confidence: 99%

Origin of the low thermal isomerization rate of rhodopsin chromophore

Yanagawa

Kojima

Yamashita

et al. 2015

Sci Rep

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“…This studies claimed that overexpression of the GRM8 gene might be used as indicator in schizophrenia pathogenesis . [24][25] Another down-regulated gene in active group was HTR2A that was shown to express serotonin receptor with various important roles. If there were abnormalities in the HTR2A, some pathogenesis could take place for instance obsessive-compulsive disorder, susceptibility to schizophrenia and major depressive disorder.…”

Section: Discussionmentioning

confidence: 99%

Comparative Expression Analysis of Genes Encoding Neurotransmitter Receptors Between Entrepreneurs and Non-Entrepreneurs

Yıldırım¹,

Aslan²,

Daştan³

et al. 2021

PJMHS

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Introduction and Purpose: Recent studies revealed that different behavioral and physiological processes are closely related to different receptor levels in humans. On the other hand, there weren’t any studies that investigated neurotransmitter activity and entrepreneurship relationships. Little is known about how genes expressed in the blood are associated with entrepreneurship. In this study, we measured the expression of 84 genes encoding neurotransmitter receptors between the entrepreneurs and non-entrepreneurs (n=25) using RT-PCR arrays to monitor differentially expressed genes for exploring molecular mechanism behind the entrepreneurship mindset. Materials and Methods: After determining whether participants are entrepreneurs or not, blood samples were collected. Blood samples were collected in Vacutainer® EDTA tubes as 10 ml and RNA isolation was performed at the Erzurum Technical University, Cell Culture Laboratory. Trizol® solution for RNA isolation (Thermo®, USA) was applied according to the manufacturer's instructions. Concentrations of RNA samples were measured at 260 nm using a spectrophotometer (Epoch®, Biotek). Then, total RNA was reverse transcribed into cDNA by using High-Capacity cDNA Reverse Transcription Kit (Thermo®). Total 84 genes were analyzed via RT² Profiler™ Human Neurotransmitter Receptors PCR Array (Qiagen®, USA) in Rotor-Gene Q real-time PCR cycler (Qiagen®, USA). Findings: The gene expression results obtained from RT-PCR were compared between entrepreneurs and non-entrepreneurs, and presented as Fold Changes (FC). According to our results, positive FC values indicated an increase in the expression of the genes and negative FC values indicated decrease in gene expression levels in entrepreneurs. Results: These 84 different genes regulate neurotransmitter biosynthesis, uptake, transport, and signaling via neurotransmitter receptors. According to gene expression analyses, gene expressions that could be related to the entrepreneur behavior might be connected to not only undesired psychological outcomes like various addictions and also neurological cases such as schizophrenia and depressive disorder. Our results firstly indicated that entrepreneurship was not only associated with neurotransmitter release but also with receptor levels. Keywords: Entrepreneurship, Entrepreneurs, Genes, Neurotransmitter Receptors, RT-PCR Array

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“…Heterologous expression of mGluR3 in HEK293 cells for in vitro biochemical assay was performed according to previously reported methods (20). The plasmid DNA (10 μ g/100-mm dish) of mGluR3 or pCAG vector (mock) was transfected into HEK293 cells growing to ~40% confluency in DMEM/F12 supplemented with 10% FBS by the calcium-phosphate method.…”

Section: Heterologous Expression and Membrane Preparation For In Vitrmentioning

confidence: 99%

“…The ECDs function as an obligatory dimer, where dimeric reorientation occurs upon ligand binding (14,15). The conformational change in ECDs promotes the dimeric rearrangement of TMDs (16)(17)(18), activating a protomer of the TMD dimer (19,20). Single-molecule tracking (SMT) analysis demonstrated that the average diffusion coefficient (D Av ) of mGluR3 quantitatively reflects receptor activity.…”

Section: Introductionmentioning

confidence: 99%

Title: Single-molecule diffusion-based estimation of ligand effects on G protein-coupled receptors

Yanagawa

Hiroshima²,

Togashi

et al. 2017

Preprint

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G protein-coupled receptors (GPCRs) are major drug targets and have high potential for drug discovery. The development of a method for measuring the activities of GPCRs is essential for pharmacology and drug screening. However, it is difficult to measure the effects of a drug by monitoring the receptor on the cell surface, and changes in the concentrations of downstream signaling molecules, which are specific to each receptor, are used as an index of the receptor activity. Here, we show that singlemolecule imaging analysis provides an alternative method for assessing GPCR activity.We monitored the dynamics of the diffusion of metabotropic glutamate receptor 3 (mGluR3), a class C GPCR, under various ligand conditions by using total internal reflection fluorescence microscopy (TIRFM). The single-molecule tracking analysis demonstrate that changes in the average diffusion coefficient of mGluR3 quantitatively reflect the ligand-dependent activity. Then, we reveal that the diffusion of receptor molecules is altered by the common physiological events associated with GPCRs, including G protein binding or accumulation in clathrin-coated pits, by inhibitor assay and dual-color single-molecule imaging analysis. We also confirm the general applicability of the method to class A and B GPCRs, demonstrating that the diffusion coefficient is a good index for estimating the activities of many GPCRs regardless of the phylogenetic groups, chemical properties of the ligands, and G protein-coupling selectivity.

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Glutamate Acts as a Partial Inverse Agonist to Metabotropic Glutamate Receptor with a Single Amino Acid Mutation in the Transmembrane Domain

Cited by 13 publications

References 36 publications

Origin of the low thermal isomerization rate of rhodopsin chromophore

Origin of the low thermal isomerization rate of rhodopsin chromophore

Comparative Expression Analysis of Genes Encoding Neurotransmitter Receptors Between Entrepreneurs and Non-Entrepreneurs

Title: Single-molecule diffusion-based estimation of ligand effects on G protein-coupled receptors

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