D-myo-inositol 1-phosphate as a surrogate of D-myo-inositol 1,4,5-tris phosphate to monitor G protein-coupled receptor activation

Autor: Audrey Michaud, Françoise Chrétien, Cyril Goudet, Jean-Philippe Pin, Hervé Ansanay, Florence Grillet, Cédric Leroy, Yves Chapleur, Fanny Malhaire, Gérard Mathis, Fabrice Maurin, Emmanuel Bourrier, Olivier Hernout, Damien Maurel, Michel Fink, Thierry Durroux, Magali Naval, Eric Trinquet, Herve Bazin
Přispěvatelé: Homogeneous Time-resolved Fluorescence (HTRF), Cis Bio International, Institut de Génomique Fonctionnelle (IGF), Université de Montpellier (UM)-Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS), Département de Pharmacologie Moléculaire, Université Montpellier 2 - Sciences et Techniques (UM2), Structure et Réactivité des Systèmes Moléculaires Complexes (SRSMC), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Université Montpellier 1 (UM1)-Université Montpellier 2 - Sciences et Techniques (UM2)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Goudet, Cyril
Jazyk: angličtina
Rok vydání: 2006
Předmět:
Tris
[SDV.BIO]Life Sciences [q-bio]/Biotechnology
Inositol Phosphates
Biophysics
Inositol monophosphatase
MESH: Cricetinae
CHO Cells
Inositol 1
4
5-Trisphosphate
MESH: Receptors
G-Protein-Coupled
Biochemistry
Receptors
G-Protein-Coupled
03 medical and health sciences
chemistry.chemical_compound
0302 clinical medicine
MESH: CHO Cells
Cricetinae
Animals
Humans
Inverse agonist
Inositol
MESH: Animals
[SDV.NEU] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]
[INFO.INFO-BT]Computer Science [cs]/Biotechnology
Inositol phosphate
Molecular Biology
030304 developmental biology
G protein-coupled receptor
chemistry.chemical_classification
0303 health sciences
MESH: Humans
biology
Cell Biology
[SDV.SP]Life Sciences [q-bio]/Pharmaceutical sciences
MESH: Inositol Phosphates
[SDV.BIO] Life Sciences [q-bio]/Biotechnology
[SDV.SP] Life Sciences [q-bio]/Pharmaceutical sciences
Enzyme
[INFO.INFO-BT] Computer Science [cs]/Biotechnology
chemistry
Type C Phospholipases
biology.protein
MESH: Inositol 1
4
5-Trisphosphate
[SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]
Signal transduction
MESH: Type C Phospholipases
030217 neurology & neurosurgery
Zdroj: Analytical Biochemistry
Analytical Biochemistry, Elsevier Masson, 2006, 358 (1), pp.126-35. ⟨10.1016/j.ab.2006.08.002⟩
Analytical Biochemistry, 2006, 358 (1), pp.126-35. ⟨10.1016/j.ab.2006.08.002⟩
ISSN: 0003-2697
1096-0309
DOI: 10.1016/j.ab.2006.08.002⟩
Popis: International audience; Phospholipase C beta (PLC-beta)-coupled G protein-coupled receptor (GPCR) activities traditionally are assessed by measuring Ca2+ triggered by D-myo-inositol 1,4,5-trisphosphate (IP3), a PLC-beta hydrolysis product, or by measuring the production of inositol phosphate using cumbersome radioactive assays. A specific detection of IP3 production was also established using IP3 binding proteins. The short lifetime of IP3 makes this detection very challenging in measuring GPCR responses. Indeed, this IP3 rapidly enters the metabolic inositol phosphate cascade. It has been known for decades that lithium chloride (LiCl) leads to D-myo-inositol 1-phosphate accumulation on GPCR activation by inhibiting inositol monophosphatase, the final enzyme of the IP3 metabolic cascade. We show here that IP1 can be used as a surrogate of IP3 to monitor GPCR activation. We developed a novel homogeneous time-resolved fluorescence (HTRF) assay that correlates perfectly with existing methods and is easily amenable to high-throughput screening. The IP-One assay was validated on various GPCR models. It has the advantage over the traditional Ca2+ assay of allowing the measurement of inverse agonist activity as well as the analysis of PLC-beta activity in any nontransfected primary cultures. Finally, the high assay specificity for D-myo-inositol 1 monophosphate (IP1(1)) opens new possibilities in developing selective assays to study the functional roles of the various isoforms of inositol phosphates.
Databáze: OpenAIRE
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