Receptor Activity-Modifying Protein 2 (RAMP2) alters glucagon receptor trafficking in hepatocytes with functional effects on receptor signalling

Pancreatic bi-hormones insulin and glucagon are the Yin and Yang in the regulation of glucose metabolism and homoeostasis. Insulin is synthesized primarily by pancreatic β-cells and is released in response to an increase in blood glucose levels (hyperglycaemia). By contrast, glucagon is synthesized by pancreatic α-cells and is released in response to a decrease in blood glucose (hypoglycaemia). The principal role of glucagon is to counter the actions of insulin on blood glucose homoeostasis, but it also has diverse non-hyperglycaemic actions. Although Type 1 diabetes is caused by insulin deficiency (insulin-dependent) and can be corrected by insulin replacement, Type 2 diabetes is a multifactorial disease and its treatment is not dependent on insulin therapy alone. Type 2 diabetes in humans is characterized by increased insulin resistance, increased fasting blood glucose, impaired glucose tolerance and the development of glomerular hyperfiltration and microalbuminuria, ultimately leading to diabetic nephropathy and end-stage renal disease. Clinical studies have suggested that an inappropriate increase in hyperglycaemic glucagon (hyperglucagonaemia) over hypoglycaemic insulin (not insulin deficiency until advanced stages) plays an important role in the pathogenesis of Type 2 diabetes. However, for decades, research efforts and resources have been devoted overwhelmingly to studying the role of insulin and insulin-replacement therapy. By contrast, the implication of glucagon and its receptor signalling in the development of Type 2 diabetic metabolic syndromes and end-organ injury has received little attention. The aim of this review is to examine the evidence as to whether glucagon and its receptor signalling play any role(s) in the pathogenesis of Type 2 diabetic renal injury, and to explore whether targeting glucagon receptor signalling remains only a theoretical antidiabetic strategy in Type 2 diabetes or may realize its promise in the future.

Download Full-text

Memory Retrieval Requires Ongoing Protein Synthesis and NMDA Receptor Activity-Mediated AMPA Receptor Trafficking

Journal of Neuroscience ◽

10.1523/jneurosci.0735-14.2015 ◽

2015 ◽

Vol 35 (6) ◽

pp. 2465-2475 ◽

Cited By ~ 48

Author(s):

J. Lopez ◽

K. Gamache ◽

R. Schneider ◽

K. Nader

Keyword(s):

Protein Synthesis ◽

Nmda Receptor ◽

Ampa Receptor ◽

Memory Retrieval ◽

Receptor Trafficking ◽

Receptor Activity ◽

Ampa Receptor Trafficking

Download Full-text

Receptor Activity-Modifying Protein 2 (RAMP2) alters glucagon receptor trafficking in hepatocytes with functional effects on receptor signalling

10.1101/2021.05.09.443291 ◽

2021 ◽

Author(s):

Emma R McGlone ◽

Yusman Manchanda ◽

Ben Jones ◽

Phil Pickford ◽

Asuka Inoue ◽

...

Keyword(s):

Plasma Membrane ◽

Viral Vector ◽

Radioligand Binding ◽

Receptor Activity ◽

Spatiotemporal Pattern ◽

Subcellular Localisation ◽

Glucagon Receptor ◽

Agonist Stimulation ◽

Huh7 Cells

Objectives: Receptor Activity-Modifying Protein 2 (RAMP2) is a chaperone protein which allosterically binds to and interacts with the glucagon receptor (GCGR). The aims of this study were to investigate the effects of RAMP2 on GCGR trafficking and signalling in the liver, where glucagon is important for carbohydrate and lipid metabolism. Methods: Subcellular localisation of GCGR in the presence and absence of RAMP2 was investigated using confocal microscopy, trafficking assays and radioligand binding assays in human embryonic kidney (HEK293T) and human hepatoma (Huh7) cells. Mouse embryonic fibroblasts (MEFs) lacking Wiskott Aldrich Syndrome protein and scar homologue (WASH) complex were used to investigate the effect of a halt in recycling of internalised proteins on GCGR signalling in the absence of RAMP2. NanoBiT complementation and cyclic AMP assays were used to study the functional effect of RAMP2 on recruitment and activation of GCGR signalling mediators. Response to hepatic RAMP2 up-regulation in lean and obese adult mice using a bespoke adeno-associated viral vector was also studied. Results: GCGR is predominantly localised at the plasma membrane in the absence of RAMP2 and exhibits remarkably slow internalisation in response to agonist stimulation. Rapid intracellular retention of glucagon-stimulated GCGR in cells lacking WASH complex indicates that activated GCGRs undergo continuous cycles of internalisation and recycling despite apparent GCGR plasma membrane localisation up to 40 minutes post-stimulation. Co-expression of RAMP2 induces GCGR internalisation both basally and in response to agonist-stimulation. The intracellular retention of GCGR in the presence of RAMP2 confers a bias away from β-arrestin-2 recruitment coupled to increased activation of Gαs proteins at endosomes. This is associated with increased short-term efficacy for glucagon-stimulated cAMP production, although long-term signalling is dampened by increased receptor lysosomal targeting for degradation. Despite these signalling effects, only minor disturbance of carbohydrate metabolism was observed in mice with up-regulated hepatic RAMP2. Conclusions: By retaining GCGR intracellularly, RAMP2 alters the spatiotemporal pattern of GCGR signalling. Further exploration of the effects of RAMP2 on GCGR in vivo is warranted.

Download Full-text

A glucagon analogue decreases body weight in mice via signalling in the liver

Scientific Reports ◽

10.1038/s41598-021-01912-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Charlotte E. Hinds ◽

Bryn M. Owen ◽

David C. D. Hope ◽

Philip Pickford ◽

Ben Jones ◽

...

Keyword(s):

Metabolic Syndrome ◽

Weight Loss ◽

Body Weight ◽

Side Effects ◽

Interindividual Variation ◽

Receptor Specificity ◽

Glucagon Receptor ◽

Receptor Signalling ◽

Long Acting

AbstractGlucagon receptor agonists show promise as components of next generation metabolic syndrome pharmacotherapies. However, the biology of glucagon action is complex, controversial, and likely context dependent. As such, a better understanding of chronic glucagon receptor (GCGR) agonism is essential to identify and mitigate potential clinical side-effects. Herein we present a novel, long-acting glucagon analogue (GCG104) with high receptor-specificity and potent in vivo action. It has allowed us to make two important observations about the biology of sustained GCGR agonism. First, it causes weight loss in mice by direct receptor signalling at the level of the liver. Second, subtle changes in GCG104-sensitivity, possibly due to interindividual variation, may be sufficient to alter its effects on metabolic parameters. Together, these findings confirm the liver as a principal target for glucagon-mediated weight loss and provide new insights into the biology of glucagon analogues.

Download Full-text