Distinct regions in the C-Terminus required for GLP-1R cell surface expression, activity and internalisation

Thompson, Aiysha; Kanamarlapudi, Venkateswarlu

doi:10.1016/j.mce.2015.06.012

Cited by 22 publications

(15 citation statements)

References 52 publications

(68 reference statements)

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“…Results are expressed as the mean + standard error of the mean (SEM) of three to five experiments. Statistical significance between multiple groups was determined by the Bonferroni's post hoc test after one-way or two-way analysis of variance (ANOVA), where p <0.05 was considered as statistically significant (Thompson and Kanamarlapudi, 2015b).…”

Section: Discussionmentioning

confidence: 99%

Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active

Kanamarlapudi

Gordon

Bernal

2016

Reproductive BioMedicine Online

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General rightsThis document is made available in accordance with publisher policies. Please cite only the published version using the reference above. AbstractPolycystic ovarian syndrome (PCOS) is associated with anovulatory infertility. Luteinizing hormone/chorionic gonadotropin receptor (LHCGR), which is critical for ovulation, has been suggested to be expressed prematurely in the ovarian follicles of women with PCOS. The objective of this study was to analyse the expression and activity of LHCGR in ovarian granulosa cells from PCOS patients and the involvement of ARF6 small GTPase in LHCGR internalisation. Granulosa cells (GCs) isolated from follicular fluid collected during oocyte retrieval from normal women (n=19) and women with PCOS (n=17) were used to study differences in LHCGR protein expression and activity between normal and PCOS patients.LHCGR expression is up-regulated in GCs from PCOS women. LHCGR in PCOS GCs is functionally active as evidenced by increased cAMP production upon human gonadotrophin (HCG)-stimulation. Moreover, ARF6 is highly expressed in GCs from PCOS patients and HCG-stimulation increases the levels of active ARF6. The inhibition of ARF6 activation attenuates HCG-induced LHCGR internalisation in both normal and PCOS GCs, indicating that there are no alterations in LHCGR internalisation in GCs from PCOS. In conclusion, the expression and activation of LHCGR and ARF6 are up-regulated in GCs from PCOS women but the mechanism of agonist-induced LHCGR internalisation is unaltered.

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Section: Discussionmentioning

confidence: 99%

Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active

Kanamarlapudi

Gordon

Bernal

2016

Reproductive BioMedicine Online

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“…The C-terminus has been repeatedly demonstrated to be an indispensable region of GPCRs that govern their cell surface expression (6). In the present study, the effect of C-terminal truncation on the surface expression of OX2R was assessed using an ELISA.…”

Section: Resultsmentioning

confidence: 99%

“…The C-termini of GPCRs differ in terms of their length and structure, implying their functional importance and complexity (5). The C-terminus is indispensable for the functions of GPCRs, including cell surface localization (6), G-protein coupling (7), agonist-driven internalization (8) and signal transduction (9). Therefore, mutations in the C-termini of GPCRs have been associated with a variety of dysfunctions and disease states (10,11).…”

Section: Introductionmentioning

confidence: 99%

C-terminus of OX2R significantly affects downstream signaling pathways

Wang

Liu

et al. 2017

Molecular Medicine Reports

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The human orexin 2 receptor (OX2R) is a G-protein-coupled receptor (GPCR) that has been implicated in a number of diverse physiological functions. Recent studies have identified a number of functions of the C-termini of GPCRs. However, the importance of the OX2R C-terminus in regulating signaling and surface expression remains unclear. In the present study, the function of the OX2R C-terminus was investigated using three C-terminal mutants, which were truncated at residues 368, 384 and 414, respectively, and the wild-type control, which expressed the full-length OX2R. HEK-293 cells were transfected with the mutated and control OX2R constructs. ELISA, western blot analysis and calcium assays were used to investigate the effects of the mutations on OX2R function. The present results demonstrated that residues 385–414 and 415–444 exhibited a cumulative effect on the surface expression of OX2R. Residues 369–384 exhibited a significant influence on inositol phosphate production and extracellular signal-regulated kinase 1/2 phosphorylation. Residues 385–414 significantly influenced agonist-induced internalization, whereas residues 369–384 and 385–414 significantly influenced Ca2+ release. The results of the present study suggest that the C-terminus of OX2R is important for its role in various physiological and pathological processes, and may therefore be associated with such disorders as depression and anorexia.

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“…We have recently shown that agonist-induced GLP-1R internalisation is mediated by the Gα q pathway [ 20 ]. In addition, the C-terminus of GLP-1R plays an important role in agonist-induced internalisation of the receptor [ 22 , 23 ].…”

Section: Introductionmentioning

confidence: 99%

Molecular Characterisation of Small Molecule Agonists Effect on the Human Glucagon Like Peptide-1 Receptor Internalisation

et al. 2016

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The glucagon-like peptide receptor (GLP-1R), which is a G-protein coupled receptor (GPCR), signals through both Gαs and Gαq coupled pathways and ERK phosphorylation to stimulate insulin secretion. The aim of this study was to determine molecular details of the effect of small molecule agonists, compounds 2 and B, on GLP-1R mediated cAMP production, intracellular Ca2+ accumulation, ERK phosphorylation and its internalisation. In human GLP-1R (hGLP-1R) expressing cells, compounds 2 and B induced cAMP production but caused no intracellular Ca2+ accumulation, ERK phosphorylation or hGLP-1R internalisation. GLP-1 antagonists Ex(9–39) and JANT-4 and the orthosteric binding site mutation (V36A) in hGLP-1R failed to inhibit compounds 2 and B induced cAMP production, confirming that their binding site distinct from the GLP-1 binding site on GLP-1R. However, K334A mutation of hGLP-1R, which affects Gαs coupling, inhibited GLP-1 as well as compounds 2 and B induced cAMP production, indicating that GLP-1, compounds 2 and B binding induce similar conformational changes in the GLP-1R for Gαs coupling. Additionally, compound 2 or B binding to the hGLP-1R had significantly reduced GLP-1 induced intracellular Ca2+ accumulation, ERK phosphorylation and hGLP-1R internalisation. This study illustrates pharmacology of differential activation of GLP-1R by GLP-1 and compounds 2 and B.

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Distinct regions in the C-Terminus required for GLP-1R cell surface expression, activity and internalisation

Cited by 22 publications

References 52 publications

Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active

Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active

C-terminus of OX2R significantly affects downstream signaling pathways

Molecular Characterisation of Small Molecule Agonists Effect on the Human Glucagon Like Peptide-1 Receptor Internalisation

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