title: Phospholipase C-ε links Epac2 activation to the potentiation of glucose-stimulated insulin secretion from mouse islets of Langerhans.
abstract: Glucose-stimulated insulin secretion (GSIS) from pancreatic β-cells is potentiated by cAMP-elevating agents, such as the incretin hormone glucagon-like peptide-1 (GLP-1), and cAMP exerts its insulin secretagogue action by activating both protein kinase A (PKA) and the cAMP-regulated guanine nucleotide exchange factor designated as Epac2. Although prior studies of mouse islets demonstrated that Epac2 acts via Rap1 GTPase to potentiate GSIS, it is not understood which downstream targets of Rap1 promote the exocytosis of insulin. Here, we measured insulin secretion stimulated by a cAMP analog that is a selective activator of Epac proteins in order to demonstrate that a Rap1-regulated phospholipase C-epsilon (PLC-ε) links Epac2 activation to the potentiation of GSIS. Our analysis demonstrates that the Epac activator 8-pCPT-2'-O-Me-cAMP-AM potentiates GSIS from the islets of wild-type (WT) mice, whereas it has a greatly reduced insulin secretagogue action in the islets of Epac2 (-/-) and PLC-ε (-/-) knockout (KO) mice. Importantly, the insulin secretagogue action of 8-pCPT-2'-O-Me-cAMP-AM in WT mouse islets cannot be explained by an unexpected action of this cAMP analog to activate PKA, as verified through the use of a FRET-based A-kinase activity reporter (AKAR3) that reports PKA activation. Since the KO of PLC-ε disrupts the ability of 8-pCPT-2'-O-Me-cAMP-AM to potentiate GSIS, while also disrupting its ability to stimulate an increase of β-cell [Ca2+]i, the available evidence indicates that it is a Rap1-regulated PLC-ε that links Epac2 activation to Ca2+-dependent exocytosis of insulin.
authors.affiliation: Department of Medicine, State University of New York, Upstate Medical University, Syracuse, NY, USA.
authors.forename: Igor
authors.initials: I
authors.lastname: Dzhura
authors.forename: Oleg G
authors.initials: OG
authors.lastname: Chepurny
authors.forename: Colin A
authors.initials: CA
authors.lastname: Leech
authors.forename: Michael W
authors.initials: MW
authors.lastname: Roe
authors.forename: Elvira
authors.initials: E
authors.lastname: Dzhura
authors.forename: Xin
authors.initials: X
authors.lastname: Xu
authors.forename: Youming
authors.initials: Y
authors.lastname: Lu
authors.forename: Frank
authors.initials: F
authors.lastname: Schwede
authors.forename: Hans-G
authors.initials: HG
authors.lastname: Genieser
authors.forename: Alan V
authors.initials: AV
authors.lastname: Smrcka
authors.forename: George G
authors.initials: GG
authors.lastname: Holz
mesh_terms: D000818:Animals; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology; D031541:Fluorescence Resonance Energy Transfer; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism / Q000502:physiology*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000201:enzymology / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D054801:Phosphoinositide Phospholipase C / Q000378:metabolism*
chemical_list: C473217:8-(4-chloro-phenylthio)-2'-O-methyladenosine-3'-5'-cyclic monophosphate; D020662:Guanine Nucleotide Exchange Factors; D007328:Insulin; C486707:Rapgef4 protein, mouse; D000242:Cyclic AMP; D054801:Phosphoinositide Phospholipase C; C423320:phospholipase C epsilon