title: Intracellular Ca(2+) oscillations generated via the extracellular Ca(2+)-sensing receptor (CaSR) in response to extracellular Ca(2+) or L-phenylalanine: Impact of the highly conservative mutation Ser170Thr.
abstract: The extracellular Ca(2+)-sensing receptor (CaSR) is an allosteric protein that responds to changes in the extracellular concentration of Ca(2+) ([Ca(2+)]e) and aromatic amino acids with the production of different patterns of oscillations in intracellular Ca(2+) concentration ([Ca(2+)]i). An increase in [Ca(2+)]e stimulates sinusoidal oscillations in [Ca(2+)]i whereas aromatic amino acid-induced CaR activation in the presence of a threshold [Ca(2+)]e promotes transient oscillations in [Ca(2+)]i. Here, we examined spontaneous and ligand-evoked [Ca(2+)]i oscillations in single HEK-293 cells transfected with the wild type CaSR or with a mutant CaSR in which Ser170 was converted to Thr (CaSRS170T). Our analysis demonstrates that cells expressing CaSRS170T display [Ca(2+)]i oscillations in the presence of low concentrations of extracellular Ca(2+) and respond to L-Phe with robust transient [Ca(2+)]i oscillations. Our results indicate that the S170T mutation induces a marked increase in CaSR sensitivity to [Ca(2+)]e and imply that the allosteric regulation of the CaSR by aromatic amino acids is not only mediated by an heterotropic positive effect on Ca(2+) binding cooperativity but, as biased agonists, aromatic amino acids stabilize a CaSR conformation that couples to a different signaling pathway leading to transient [Ca(2+)]i oscillations.
authors.affiliation: Division of Digestive Diseases, Department of Medicine, CURE: Digestive Diseases Research Center, David Geffen School of Medicine and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, CA 90095, USA.
authors.forename: Steven H
authors.initials: SH
authors.lastname: Young
authors.affiliation: Institute of Immunology, Genetics and Metabolism, Consejo Nacional de Investigaciones Cientificas y Tecnicas, Universidad de Buenos Aires, Buenos Aires, Argentina.
authors.forename: Osvaldo
authors.initials: O
authors.lastname: Rey
authors.affiliation: Division of Digestive Diseases, Department of Medicine, CURE: Digestive Diseases Research Center, David Geffen School of Medicine and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, CA 90095, USA; Veterans Affairs West Los Angeles Healthcare System, Los Angeles, CA, USA. Electronic address: erozengurt@mednet.ucla.edu.
authors.forename: Enrique
authors.initials: E
authors.lastname: Rozengurt
mesh_terms: D019943:Amino Acid Substitution; D001665:Binding Sites / Q000235:genetics; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010649:Phenylalanine / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D044169:Receptors, Calcium-Sensing / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: C095550:CASR protein, human; D050505:Mutant Proteins; D044169:Receptors, Calcium-Sensing; D011994:Recombinant Proteins; D010649:Phenylalanine; D002118:Calcium
keywords: Allosteric regulation; Amino acid signaling; Ca(2+) oscillations; CaSR mutant S170T