The third cytoplasmic domain of the GLP-1[7-36 amide] receptor is required for coupling to the adenylyl cyclase system.

Takhar, Sarvjinder; Gyomorey, S.; Su, Ruey Chyi; Mathi, Swarna K.; Li, Xinfang; Wheeler, Michael B.

doi:10.1210/endo.137.5.8612565

Cited by 56 publications

(17 citation statements)

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“…These data are consistent with the results of the human m 1 muscarinic receptor (36). In contrast, deletion of the threonine, aspartic acid, and isoleucine (TDI) motif within the central region of the third endoloop of the GLP-1 receptor led to a significant reduction in receptor expression (30). The corresponding mutant in the hSR is IC3-3 with the glycine, asparagine, glutamic acid, valine (GNEV) motif deleted, and this mutant is functionally indistinguishable from the WT receptor.…”

Section: Discussionsupporting

confidence: 90%

“…Within the third endoloop, the function of the conserved basic motif at the N-terminal region as the site for G protein interaction has been a focus of interest, and the results obtained are controversial. Evidence supporting this notion can be found in studies of the GLP-1, rhodopsin, and ␤ 2-adrenergic and muscarinic receptors (30,(32)(33)(34). Instead, other reports have suggested that the hydrophobic residues at the junction of the fifth transmembrane domain of GLP-1 receptor (29) and those at the end of the third endoloop of ␤-adrenergic receptor (35) are involved in G protein coupling.…”

Section: Discussionmentioning

confidence: 80%

“…coupling of G protein (30). As yet, there is no evidence to implicate the role of the RLAR motif in G protein activation in the secretin receptor family.…”

mentioning

confidence: 98%

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Functional Segregation of the Highly Conserved Basic Motifs within the Third Endoloop of the Human Secretin Receptor

2001

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In this study, a mutagenesis-based strategy was employed to assess the roles of two highly conserved motifs (KLR and RLAR) within the third endoloop of the human secretin receptor. Block deletion of KLRT and mutation of Lys323 (K 323 I) significantly reduced cAMP accumulation, and these mutations did not affect ligand interaction and receptor number expressed on the cell surface. Thus, the KLRT region at the N terminus of the third endoloop, particularly Lys323, is important for G protein coupling. For the RLAR motif, receptors with substitutions at positions 339 and 342 from Arg to Ala (R 339, 342 A), Glu (R 339, 342 E), or Ile (R 339, 342 I) as well as block deletion of the RLAR motif were all found to be defective in both secretin-binding and cAMP production. Interestingly, a single mutation at the corresponding positions of Arg339 or Arg342 responded as the wild-type human secretin receptor in all functional assays, indicating that the presence of one Arg at either position within the RLAR motif is sufficient for a normal receptor function. Immunofluorescent staining of these mutant receptors showed that these Arg residues are responsible for surface presentation and/or receptor stability. (Endocrinology 142: 3926 -3934, 2001)T HE SECRETIN RECEPTOR has a high affinity for secretin and a relatively low affinity for VIP (1) and belongs to the class II G protein-coupled receptor (GPCR) subfamily. This secretin/VIP receptor subfamily also includes receptors for glucagon, glucagon-like peptide-1 (GLP-1), gastric inhibitory peptide, PTH, pituitary adenylate cyclase activating polypeptide, and GHRH. Generally, the signal transduction mechanism of GPCRs involves ligandinduced changes that affect the conformation of the intracellular surface of the receptors and hence promote the coupling to G proteins (2-6). In the case of the secretin/VIP receptor subfamily, this stimulation process activates adenylate cyclase and eventually leads to the elevation of intracellular cAMP (7-11). Besides cAMP, other intracellular second messengers, such as Ca 2ϩ and inositol phosphates have also been reported (12)(13)(14).Like other GPCRs, the secretin receptor displays a common structural profile in which seven transmembrane domains are linked by alternative exo-and endoloops. Within the same class, there are many conserved amino acid residues, including six well-conserved cysteine residues in the N-terminal extracellular domain and multiple consensus N-glycosylation sites. Nevertheless, these receptors share only 25-50% of amino acid identity among themselves. When compared with other classes of receptors, such as the rhodopsin/␤-adrenergic receptor family, the secretin receptor family is distinct with respect to the primary sequence. Even with this minimal level of sequence homology, comparisons of receptor properties within the family can provide insight into the importance of specific structural domains or motifs. For example, the diversity of the N termini of these receptors in amino acid composition suggests that the N-terminal ...

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Section: Discussionsupporting

confidence: 90%

Section: Discussionmentioning

confidence: 80%

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Functional Segregation of the Highly Conserved Basic Motifs within the Third Endoloop of the Human Secretin Receptor

2001

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“…Previous studies with the rat GLP-1 receptor demonstrated that the KLK motif, and in particular the first lysine residue, is required for the efficient coupling of the receptor to adenylyl cyclase. Alteration of this motif or the lysine residue in the GLP-1 receptor resulted in significantly reduced cAMP production (59). The KLR motif of the glucagon receptors is found in the same position as the KLK motif of GLP-1 receptors (Fig.…”

Section: Discussionmentioning

confidence: 83%

Identification and Characterization of a Glucagon Receptor from the Goldfish Carassius auratus: Implications for the Evolution of the Ligand Specificity of Glucagon Receptors in Vertebrates

et al. 2004

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The structural basis of ligand selectivity of G protein-coupled receptors for metabolic hormones has been an area of intense investigation, and yet it remains unresolved. One approach to delineating the mechanism of ligand-receptor interactions is to compare the ligand specificities of receptors expressed in species that emerged at different times within vertebrate evolution. In this paper we describe the isolation, functional, and phylogenetic characterization of the glucagon receptor from the goldfish Carassius auratus (Teleostei, order Cypriniformes), and compare its ligand specificity with that of the homologous rat receptor. Goldfish (gf) glucagon stimulated glucose production in a dose-dependent manner from isolated goldfish hepatocytes, resulting in 5-fold increase at 1 microm. The goldfish glucagon receptor (gfGlucR) shares 56, 51, 50, and 52% amino acid identities with frog Rana tigrina regulosa, mouse, rat, and human glucagon receptors, respectively. In competitive binding experiments, the recombinant gfGlucR displays high affinity toward goldfish, zebrafish, and human glucagons (IC(50) = 0.6, 9, and 13 nm, respectively) but not toward goldfish glucagon-like peptide-1 or human glucagon-like peptide-1 (7-36) amide. Whereas both goldfish and human glucagons stimulated dose-dependent increases in intracellular cAMP through the recombinant gfGlucR, the recombinant rat GlucR interacted only with human glucagon, analogous to the specificity of the previously characterized glucagon receptor from the frog R. tigrina regulosa. Our results demonstrate that the binding pocket of gfGlucR can accommodate a broad range of glucagon structures and that in the frogs and mammals, there is a structural switch to a more restrictive conformation of glucagon receptors.

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“…Within the third intracellular loop, mutation of a central lysine-leucine motif of the opossum parathyroid hormone receptor, rat GLP1 receptor, and human VPAC1 and VPAC2 receptors, analogous to the human secretin receptor residues Lys 302 -Leu 303 (ICL3A), results in defects in agonist-stimulated cAMP production, calcium mobilization, and ligand binding to varying degrees (1,7,25,38,45,62). In the present work, replacing these residues with alanines in the human secretin receptor resulted in normal ligand binding and calcium signaling, but reduced cAMP responses.…”

Section: Discussionmentioning

confidence: 99%

Differential determinants for coupling of distinct G proteins with the class B secretin receptor

Garcia

Dong

Miller

2012

American Journal of Physiology-Cell Physiology

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The secretin receptor is a prototypic class B G protein-coupled receptor that is activated by binding of its natural peptide ligand. The signaling effects of this receptor are mediated by coupling with Gs, which activates cAMP production, and Gq, which activates intracellular calcium mobilization. We have explored the molecular basis for the coupling of each of these G proteins to this receptor using systematic site-directed mutagenesis of key residues within each of the intracellular loop regions, and studying ligand binding and secretin-stimulated cAMP and calcium responses. Mutation of a conserved histidine in the first intracellular loop (H157A and H157R) markedly reduced cell surface expression, resulting in marked reduction in cAMP and elimination of measurable calcium responses. Mutation of an arginine (R153A) in the first intracellular loop reduced calcium, but not cAMP responses. Mutation of a dibasic motif in the second intracellular loop (R231A/K232A) had no significant effects on any measured responses. Mutations in the third intracellular loop involving adjacent lysine and leucine residues (K302A/L303A) or two arginine residues separated by a leucine and an alanine (R318A/R321A) significantly reduced cAMP responses, while the latter also reduced calcium responses. Additive effects were elicited by combining the effective mutations, while combining all the effective mutations resulted in a construct that continued to bind secretin normally, but that elicited no significant cAMP or calcium responses. These data suggest that, while some receptor determinants are clearly shared, there are also distinct determinants for coupling with each of these G proteins.

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The third cytoplasmic domain of the GLP-1[7-36 amide] receptor is required for coupling to the adenylyl cyclase system.

Cited by 56 publications

References 0 publications

Functional Segregation of the Highly Conserved Basic Motifs within the Third Endoloop of the Human Secretin Receptor

Functional Segregation of the Highly Conserved Basic Motifs within the Third Endoloop of the Human Secretin Receptor

Identification and Characterization of a Glucagon Receptor from the Goldfish Carassius auratus: Implications for the Evolution of the Ligand Specificity of Glucagon Receptors in Vertebrates

Differential determinants for coupling of distinct G proteins with the class B secretin receptor

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