Human GIP(3-30)NH2 inhibits G protein-dependent as well as G protein-independent signaling and is selective for the GIP receptor with high-affinity binding to primate but not rodent GIP receptors.
Biochem Pharmacol
; 150: 97-107, 2018 04.
Article
en En
| MEDLINE
| ID: mdl-29378179
ABSTRACT
GIP(3-30)NH2 is a high affinity antagonist of the GIP receptor (GIPR) in humans inhibiting insulin secretion via G protein-dependent pathways. However, its ability to inhibit G protein-independent signaling is unknown. Here we determine its action on arrestin-recruitment and receptor internalization in recombinant cells. As GIP is adipogenic, we evaluate the inhibitory actions of GIP(3-30)NH2 in human adipocytes. Finally, we determine the receptor selectivity of GIP(3-30)NH2 among other human and animal GPCRs. cAMP accumulation and ß-arrestin 1 and 2 recruitment were studied in transiently transfected HEK293 cells and real-time internalization in transiently transfected HEK293A and in HEK293A ß-arrestin 1 and 2 knockout cells. Furthermore, human subcutaneous adipocytes were assessed for cAMP accumulation following ligand stimulation. Competition binding was examined in transiently transfected COS-7 cells using human 125I-GIP(3-30)NH2. The selectivity of human GIP(3-30)NH2 was examined by testing for agonistic and antagonistic properties on 62 human GPCRs. Human GIP(3-30)NH2 inhibited GIP(1-42)-induced cAMP and ß-arrestin 1 and 2 recruitment on the human GIPR and Schild plot analysis showed competitive antagonism with a pA2 and Hill slope of 16.8â¯nM and 1.11⯱â¯0.02 in cAMP, 10.6â¯nM and 1.15⯱â¯0.05 in ß-arrestin 1 recruitment, and 10.2â¯nM and 1.06⯱â¯0.05 in ß-arrestin 2 recruitment. Efficient internalization of the GIPR was dependent on the presence of either ß-arrestin 1 or 2. Moreover, GIP(3-30)NH2 inhibited GIP(1-42)-induced internalization in a concentration-dependent manner and notably also inhibited GIP-mediated signaling in human subcutaneous adipocytes. Finally, the antagonist was established as GIPR selective among 62 human GPCRs being species-specific with high affinity binding to the human and non-human primate (Macaca fascicularis) GIPRs, and low affinity binding to the rat and mouse GIPRs (Kd values of 2.0, 2.5, 31.6 and 100â¯nM, respectively). In conclusion, human GIP(3-30)NH2 is a selective and species-specific GIPR antagonist with broad inhibition of signaling and internalization in transfected cells as well as in human adipocytes.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Fragmentos de Péptidos
/
Receptores de la Hormona Gastrointestinal
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Polipéptido Inhibidor Gástrico
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Transducción de Señal
/
Receptores Acoplados a Proteínas G
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Biochem Pharmacol
Año:
2018
Tipo del documento:
Article
País de afiliación:
Dinamarca