scholarly journals Agonist-induced internalization and recycling of the glucagon-like peptide-1 receptor in transfected fibroblasts and in insulinomas

1995 ◽  
Vol 310 (1) ◽  
pp. 203-214 ◽  
Author(s):  
C Widmann ◽  
W Dolci ◽  
B Thorens
Keyword(s):  
Plasma Membrane ◽  
Pancreatic Beta Cell ◽  
Chinese Hamster ◽  
Lung Fibroblasts ◽  
Continuous Exposure ◽  
Agonist Binding ◽  
Receptor Endocytosis ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Insulinoma Cells

Glucagon-like peptide-1 (GLP-1) is the most potent stimulator of glucose-induced insulin secretion and its pancreatic beta-cell receptor is a member of a new subfamily of G-protein-coupled receptors which includes the receptors for vasoactive intestinal polypeptide, secretin and glucagon. Here we studied agonist-induced GLP-1 receptor internalization in receptor-transfected Chinese hamster lung fibroblasts using three different approaches. First, iodinated GLP-1 bound at 4 degrees C to transfected cells was internalized with a t 1/2 of 2-3 min following warming up of the cells to 37 degrees C. Secondly, exposure to GLP-1 induced a shift in the distribution of the receptors from plasma membrane-enriched to endosomes-enriched membrane fractions, as assessed by Western blot detection of the receptors using specific antibodies. Thirdly, continuous exposure of GLP-1 receptor-expressing cells to iodinated GLP-1 led to a linear accumulation of peptide degradation products in the medium following a lag time of 20-30 min, indicating a continuous cycling of the receptor between the plasma membrane and endosomal compartments. Potassium depletion and hypertonicity inhibited transferrin endocytosis, a process known to occur via coated pit formation, as well as GLP-1 receptor endocytosis. In contrast to GLP-1, the antagonist exendin-(9-39) did not lead to receptor endocytosis. Surface re-expression following one round of GLP-1 receptor endocytosis occurred with a half-time of about 15 min. The difference in internalization and surface re-expression rates led to a progressive redistribution of the receptor in intracellular compartments upon continuous exposure to GLP-1. Finally, endogenous GLP-1 receptors expressed by insulinoma cells were also found to be internalized upon agonist binding. Together our data demonstrate that the GLP-1 receptor is internalized upon agonist binding by a route similar to that taken by single transmembrane segment receptors. The characterization of the pathway and kinetics of GLP-1-induced receptor endocytosis will be helpful towards understanding the role of internalization and recycling in the control of signal transduction by this receptor.

scholarly journals Agonist binding affinity determines palmitoylation of the glucagon-like peptide-1 receptor and its functional interaction with plasma membrane nanodomains in pancreatic beta cells

2018 ◽  
Author(s):  
Teresa Buenaventura ◽  
William E Laughlin ◽  
Stavroula Bitsi ◽  
Thomas Burgoyne ◽  
Zekun Lyu ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Binding Affinity ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Cluster Formation ◽  
Acyl Chain ◽  
Dependent Manner ◽  
Agonist Binding ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1

The glucagon-like peptide-1 receptor (GLP-1R), a key pharmacological target in type 2 diabetes and obesity, is known to undergo palmitoylation by covalent ligation of an acyl chain to cysteine 438 in its carboxyl-terminal tail. Work with other GPCRs indicates that palmitoylation can be dynamically regulated to allow receptors to partition into plasma membrane nanodomains that act as signaling hotspots. Here, we demonstrate that the palmitoylated state of the GLP-1R is increased by agonist binding, leading to its segregation and clustering into plasma membrane signaling nanodomains before undergoing internalization in a clathrin-dependent manner. Both GLP-1R signaling and trafficking are modulated by strategies targeting nanodomain segregation and cluster formation, including depletion of cholesterol or expression of a palmitoylation-defective GLP-1R mutant. Differences in receptor binding affinity exhibited by biased GLP-1R agonists, and modulation of binding kinetics with the positive allosteric modulator BETP, influence GLP-1R palmitoylation, clustering, nanodomain signaling, and internalization. Downstream effects on insulin secretion from pancreatic beta cells indicate that these processes are relevant to GLP-1R physiological actions and might be therapeutically targetable.

Effects of Glucagon-Like Peptide-1 (GLP-1) in RINm5F Insulinoma Cells

2009 ◽  
Vol 103 (S 02) ◽  
pp. 31-36 ◽  
Author(s):  
P. M. Jehle ◽  
D. Jehle ◽  
R. D. Fußgänger ◽  
G. Adler
Keyword(s):  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Insulinoma Cells

Dexamethasone pretreatment of rat insulinoma cells decreases binding of glucagon-like peptide-1(7–36)amide

1990 ◽  
Vol 126 (3) ◽  
pp. 445-450 ◽  
Author(s):  
G. Richter ◽  
R. Göke ◽  
B. Göke ◽  
R. Arnold
Keyword(s):  
Biological Action ◽  
Inhibitory Effects ◽  
Rinm5f Cells ◽  
Receptor Number ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Insulinoma Cells ◽  
Camp Formation ◽  
Rat Insulinoma ◽  
Stimulation Of

ABSTRACT The effect of dexamethasone on binding of glucagonlike peptide-1(7–36)amide (GLP-1(7–36)amide) to rat insulinoma-derived cells (RINm5F) was investigated. Preincubation of RINm5F cells with dexamethasone (100 nmol/l) for 24 h resulted in a decrease of GLP1(7-36)amide binding to 55·0±8·16% (mean ± s.e.m.), incubation for 48 h to 39·1±1·76%, and for 72 h to 15·5±4·35% of maximal binding. The GLP-1(7–36)amide-induced stimulation of cyclic AMP (cAMP) production was significantly decreased to 61·03±7·4% of maximum production in cells pretreated with dexamethasone (100 nmol/l) for 48 h. The decreased binding was due to a reduction of the receptor number while the receptor affinity remained unchanged. These inhibitory effects on binding and cAMP formation induced by dexamethasone were completely abolished when the antiglucocorticoid RU 38486 (100 nmol/l) was added during preincubation with dexamethasone. RU 38486 alone had no effects. Our data suggest that the biological action of GLP-1(7–36) amide at the B-cell may be modified by glucocorticoids. Journal of Endocrinology (1990) 126, 445–450

scholarly journals Glucagon-Like Peptide-1 Triggers Protective Pathways in Pancreatic Beta-Cells Exposed to Glycated Serum

2013 ◽  
Vol 2013 ◽  
pp. 1-10 ◽  
Author(s):  
Alessandra Puddu ◽  
Roberta Sanguineti ◽  
Arianna Durante ◽  
Alessio Nencioni ◽  
François Mach ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Cell Function ◽  
Pancreatic Beta Cell ◽  
Redox Balance ◽  
Insulin Gene ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Pancreatic Beta Cell Function

Advanced glycation end products (AGEs) might play a pathophysiological role in the development of diabetes and its complications. AGEs negatively affect pancreatic beta-cell function and the expression of transcriptional factors regulating insulin gene. Glucagon-like peptide-1 (GLP-1), an incretin hormone that regulates glucose homeostasis, might counteract the harmful effects of AGEs on the beta cells in culture. The aim of this study was to identify the intracellular mechanisms underlying GLP-1-mediated protection from AGE-induced detrimental activities in pancreatic beta cells. HIT-T15 cells were cultured for 5 days with glycated serum (GS, consisting in a pool of AGEs), in the presence or absence of 10 nmol/L GLP-1. After evaluation of oxidative stress, we determined the expression and subcellular localization of proteins involved in maintaining redox balance and insulin gene expression, such as nuclear factor erythroid-derived 2 (Nrf2), glutathione reductase, PDX-1, and MafA. Then, we investigated proinsulin production. The results showed that GS increased oxidative stress, reduced protein expression of all investigated factors through proteasome activation, and decreased proinsulin content. Furthermore, GS reduced ability of PDX-1 and MafA to bind DNA. Coincubation with GLP-1 reversed these GS-mediated detrimental effects. In conclusion, GLP-1, protecting cells against oxidants, triggers protective intercellular pathways in HIT-T15 cells exposed to GS.

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