Cyclic AMP sensor EPAC proteins and energy homeostasis

doi:10.1016/j.tem.2013.10.004

Trends in Endocrinology & Metabolism

Volume 25, Issue 2, February 2014, Pages 60-71

https://doi.org/10.1016/j.tem.2013.10.004 Get rights and content

Highlights

•
EPAC1 desensitizes leptin signaling in hypothalamus.
•
EPAC2 facilitates GSIS in response to GLP-1 and obesity induced insulin resistance.
•
Small molecule EPAC modulators have therapeutic potential for obesity and T2DM.

The pleiotropic second-messenger cAMP plays a crucial role in mediating the effects of various hormones on metabolism. The major intracellular functions of cAMP are transduced by protein kinase A (PKA) and by exchange proteins directly activated by cAMP (EPACs). The latter act as guanine-nucleotide exchange factors for the RAS-like small G proteins Rap1 and Rap2. Although the role of PKA in regulating energy balance has been extensively studied, the impact of EPACs remains relatively enigmatic. This review summarizes recent genetic and pharmacological studies concerning EPAC involvement in glucose homeostasis and energy balance via the regulation of leptin and insulin signaling pathways. In addition, the development of small-molecule EPAC-specific modulators and their therapeutic potential for the treatment of diabetes and obesity are discussed.

Section snippets

EPAC proteins; new kids on the block

cAMP is the prototypic second messenger discovered by Earl Sutherland and colleagues to play a key role in mediating the intracellular effects of hormones [1]. The deciphering of the cAMP signaling pathway is of major historical significance in biology and has led to scientific breakthroughs including the discoveries of cAMP-dependent protein kinase/protein kinase A (PKA), G proteins, and G protein-coupled receptors (GPCRs), for which Robert Lefkowitz and Brian Kobilka shared the latest Nobel

EPAC and leptin resistance

Obesity is the result of a prolonged imbalance between energy intake and expenditure [11]. The identification of the leptin gene (ob) provided a breakthrough in our understanding of obesity at the molecular level [12]. Leptin, an appetite-suppressing hormone derived from adipose tissue, plays a key role in the central regulation of satiety and energy expenditure [13]. Leptin binds to and activates the ‘long form’ of leptin receptor (OB-Rb or LepRb), a single-transmembrane-domain cytokine

EPAC2 in insulin secretion

GSIS is the fundamental pathway of insulin release from pancreatic β cells [38] (Figure 2A). After transport into β cells by facilitated diffusion, glucose undergoes oxidative phosphorylation, yielding an increase in the ATP/ADP ratio. The rise in ATP levels in turn leads to the closure of ATP-sensitive potassium channels (K_ATP), triggering membrane depolarization and the opening of L-type voltage-dependent Ca²⁺ channels (VDCCs) which allow influx of extracellular Ca²⁺ (Figure 2C). The

Targeting EPAC for therapeutic intervention

The studies reviewed here suggest that EPAC1 and EPAC2 play important roles in regulating leptin and insulin signaling, and represent attractive drug targets for the treatment of obesity and type 2 diabetes mellitus (T2DM). Small-molecule EPAC-selective modulators have been developed (Box 4) and can be further explored for their therapeutic potential 58, 59, 60, 61. Considering that EPAC1 and EPAC2 exert diverse and distinct functions, isoform-specific EPAC modulators, particularly

Concluding remarks

In summary, the roles of EPAC1 and EPAC2 in energy balance are complex. Despite recent major advances, our understanding remains limited and controversy persists in particular cases (Box 5). The generation of additional tissue-specific and conditional EPAC-knockout animal models will help to define further the physiological functions of EPAC. At the molecular and cellular levels, the dissection of EPAC signalosomes within the context of compartmentalized cAMP signaling will lead to further

Acknowledgments

We are indebted to Dr George Holz for providing critical comments. We thank everyone who has contributed to the field of EPAC signaling and apologize to those who were not cited owing to space limitations. M.A. is a recipient of a training fellowship from the Keck Center for Interdisciplinary Bioscience Training of the Gulf Coast Consortia funded by the National Institutes of Health (NIH) National Institute of General Medicine Grant T32GM089657. X.C. is supported by NIH grants R01GM066170 and

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Citation Excerpt :
These isoforms present more or less conserved structural features, except that Epac2 is the advanced version overlapping sequence of Epac1. Both of these isoforms are expressed by independent genes(Almahariq et al., 2014; De Rooij, 1998) The gene on/off phenomena of Epac's in different organs confers them a tissue-specific response, isoform-specificity, as well as specific therapeutic importance. Based on compartmentalization, tissue distribution, and molecular interactions at various loci, these isoforms vary their functionality.
cAMP-dependent guanine nucleotide exchange factor (Epac) is a key regulator in signal transduction and represents an excellent drug target to be investigated against various diseases. To date, very few modulators selective for Epac are available; however, there is still an unmet need of isoform-selective inhibitors. In the present study, ligand-based pharmacophores were designed to investigating structurally diverse molecules as Epac2 inhibitors. Pharmacophore models were developed using reported allosteric site inhibitors. The developed models were used to screen 95 thousand compounds from the National Cancer Institute (NCI), Maybride, and our in-house ICCBS Database. The binding mode and efficiency of the screened hits was investigated using molecular docking simulation on the allosteric site of Epac2 apo-protein (PDB ID: 2BYV) followed by ADMET (Absorption, Distribution, Metabolism, Excretion, and Toxicity) profiling Furthermore, obtained in silico screened hits were subjected to in vitro assay for insulin secretion. We identified, three lead molecules RDR02145, AAK-399, and AAD-026 reducing, insulin secretion. Remarkably, a higher inhibitory effect on insulin secretion was observed in AAK-399, and AAD-026 as compared to that of standard Epac2 non-competitive allosteric site inhibitor, MAY0132. Furthermore, Dynamic simulation studies of lead compounds proved the structural stability of the Epac2 auto-inhibited state. These findings underline the potential of these compounds as valuable pharmacological tools for designing future selective probes to inhibit the Epac-mediated signaling pathway.
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Min6 cells, a mouse β cell line derived from transgenic mouse expressing the large T-antigen of SV40 in pancreatic beta cells, are commonly utilized as an in vitro cellular model for investigating targets involved in insulin secretion. Epac2, an exchange protein that can be directly activated by cyclic AMP (cAMP), is critical for pharmacologic stimuli-induced insulin secretion and has been hypothesized to be a direct target of sulfonylurea. Previous loss of function studies only specifically knocked out EPAC2 isoform A, leaving the other two isoforms intact. In this study, we investigated the function of EPAC2 in Min6 cells by generating EPAC2 knock-out sublines using CRISPR-Cas9 technology, by removing all three isoforms of EPAC2. Our results indicate that Min6 cells can be successfully cloned from a single cell after electroporation with plasmids expressing EPAC2 specific guide RNA, Cas9 and GFP, followed by sorting for GFP expressing single cells. Two clones were found to have a single nucleotide deletion in targeted site of EPAC2 gene by sequencing, therefore creating a frame shift in exon 13. The EPAC2 null clones have an unexpectedly increased secretion of insulin at basal level and an elevated total intracellular insulin content. However, EPAC2 deficiency impaires glucose and sulfonylurea induced insulin secretion without affecting sulfonylurea binding to cells. Potassium chloride induced insulin secretion remains intact. Interestingly, cAMP levels remained unchanged in EPAC2 null cells during these processes. To understand the global function of EPAC2, RNA Seq study was performed, which reveals that EPAC2 deficiency affects expression of multiple previously unrecognized genes, suggesting that EPAC2 can function through multiple pathways in addition to being a cAMP sensor.
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Trends in Endocrinology & Metabolism

ReviewCyclic AMP sensor EPAC proteins and energy homeostasis

Highlights

Section snippets

EPAC proteins; new kids on the block

EPAC and leptin resistance

EPAC2 in insulin secretion

Targeting EPAC for therapeutic intervention

Concluding remarks

Acknowledgments

J. Biol. Chem.

Cell Metab.

Cell Metab.

J. Biol. Chem.

Am. J. Pathol.

J. Biol. Chem.

Cell. Signal.

Cell. Signal.

Mol. Cell. Endocrinol.

Cell

Cell

Pharmacol. Ther.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Cell Metab.

Cell Metab.

J. Biol. Chem.

J. Biol. Chem.

Mol. Cell. Endocrinol.

Vitam. Horm.

J. Biol. Chem.

Cell. Signal.

The relationship of epinephrine and glucagon to liver phosphorylase. IV. Effect of epinephrine and glucagon on the reactivation of phosphorylase in liver homogenates

J. Biol. Chem.

Profile of Brian K. Kobilka and Robert J. Lefkowitz, 2012 Nobel laureates in chemistry

Proc. Natl. Acad. Sci. U.S.A.

Cyclic nucleotide research – still expanding after half a century

Nat. Rev. Mol. Cell Biol.

CNG and HCN channels: two peas, one pod

Annu. Rev. Physiol.

Epac is a Rap1 guanine-nucleotide-exchange factor directly activated by cyclic AMP

Nature

A family of cAMP-binding proteins that directly activate Rap1

Science

Epac and PKA: a tale of two intracellular cAMP receptors

Acta Biochim. Biophys. Sin. (Shanghai)

Exchange protein directly activated by cAMP (epac): a multidomain cAMP mediator in the regulation of diverse biological functions

Pharmacol. Rev.

Epac: defining a new mechanism for cAMP action

Annu. Rev. Pharmacol. Toxicol.

Global obesity: trends, risk factors and policy implications

Nat. Rev. Endocrinol.

Positional cloning of the mouse obese gene and its human homologue

Nature

Sixteen years and counting: an update on leptin in energy balance

J. Clin. Invest.

The full-length leptin receptor has signaling capabilities of interleukin 6-type cytokine receptors

Proc. Natl. Acad. Sci. U.S.A.

Leptin activation of Stat3 in the hypothalamus of wild-type and ob/ob mice but not db/db mice

Nat. Genet.

Intracellular signalling. Key enzyme in leptin-induced anorexia

Nature

Mechanisms of leptin action and leptin resistance

Annu. Rev. Physiol.

Recent advances in understanding leptin signaling and leptin resistance

Am. J. Physiol. Endocrinol. Metab.

The importance of dietary modulation of cAMP and insulin signaling in adipose tissue and the development of obesity

Ann. N. Y. Acad. Sci.

Genetically lean mice result from targeted disruption of the RII beta subunit of protein kinase A

Nature

Enhanced leptin sensitivity, reduced adiposity, and improved glucose homeostasis in mice lacking exchange protein directly activated by cyclic AMP isoform 1

Mol. Cell. Biol.

Unbiased identification of substrates for the Epac1-inducible E3 ubiquitin ligase component SOCS-3

Biochem. Soc. Trans.

Signal transduction pathways for leptin: an embarrassment of riches

Diabetes

PTP1B and SHP2 in POMC neurons reciprocally regulate energy balance in mice

J. Clin. Invest.

Intracellular leptin-signaling pathways in hypothalamic neurons: the emerging role of phosphatidylinositol-3 kinase–phosphodiesterase-3B–cAMP pathway

Review
Cyclic AMP sensor EPAC proteins and energy homeostasis