(R4803) Cyclic AMP-dependent Activation of ERK Via GLP-1 Receptor Signaling Requires the Neuroendocrine Cell-Selective Guanine Nucleotide Exchanger NCS-RapGEF2

Wenqin Xu (NIMH-IRP)| Sam Dahlke (NIMH-IRP)| Andrew Emery (NIMH-IRP)| Oleg Chepurny (Department of Medicine, Upstate Medical University, SUNY)| George Holz (Department of Medicine, Upstate Medical University, SUNY)| Lee Eiden (NIMH-IRP)

G-protein coupled receptors activate various intracellular processes upon ligand engagement depending on coupling to adenylyl cyclase (Gs, Gi/o), phospholipase C (Gq), or RhoGEFs (G12/13). Cyclic AMP activation of the Rap-Braf-MEK-ERK pathway after signaling initiated by the neuropeptide PACAP, via the Gs-protein coupled receptor (GsPCR) PAC1, occurs uniquely through the neuritogenic cAMP sensor Rap guanine nucleotide exchange factor 2 (NCS-RapGEF2) in NS-1 neuroendocrine cells. We examined the expression of other Family B GsPCRs in this cell line and assessed cAMP elevation and neuritogenesis after treatment with their cognate peptide ligands. Exposure of NS-1 cells to the VIPR1/2 agonist VIP, or the GLP1R agonist exendin-4, did not induce neuritogenesis, or elevation of cAMP, presumably due to insufficient receptor protein expression. VIP and exendin-4 did induce neuritogenesis after transduction of human VIPR1, VIPR2 and GLP1R into NS-1 cells. Exendin-4/GLP1R-stimulated neuritogenesis was MEK-ERK- dependent (blocked by U0126), indicating its use of the cAMP->RapGEF2->ERK neuritogenic signaling pathway previously identified for PACAP/PAC1 signaling in NS-1 cells. NCS- RapGEF2 is expressed in the beta cell tumor-derived cell lines MIN6 and INS-1, and in human pancreatic islets. As in NS-1 cells, exendin-4 caused ERK phosphorylation in INS-1 cells. Reduction in RapGEF2 expression after its specific shRNA treatment reduced exendin-4-induced ERK phosphorylation. Transcriptome analysis of INS-1 cells after one hour exposure to exendin- 4 revealed an immediate early-gene response that was composed of both ERK-dependent and ERK-independent signaling targets. ERK-dependent targets, and presumably involving NCS-RapGEF2 signaling, included Nr4a2, Btg1, Skil, Plkl2, Per1 and Dyrk3, while ERK-independent targets and presumably involving PKA or Epac signaling, included Nr4a1, Nr4a3, Crem, Fosl2, Sik1 and Rgs2. We propose that cAMP signaling initiated by GLP-1 in pancreatic beta cells causes parallel activation of multiple cAMP effectors, including NCS- RapGEF2, Epac, and protein kinase A, to separately control various facets of GLP-1 action, including insulin secretion and transcriptional modulation. Identification of coupling of GsPCRs to ERK activation via NCS-RapGEF2 provides a novel signaling mode for GLP-1 in pancreatic beta cells and potentially brain and elsewhere. RapGEF2-dependent ERK activation is likely to be a feature of additional nervous and endocrine targets of family B receptor ligands including PACAP, VIP, CRH, GIP, secretin, glucagon, PTH, calcitonin, and GHRH. Whether or not ligands for these receptors, including GLP-1, might activate the NCS-RapGEF2, PKA, or Epac cAMP-dependent pathways in a biased fashion, with differential physiological effects, has yet to be investigated.

This abstract is dedicated to the memory of Andrew C. Emery. 

Support or Funding Information

NIMHIRPMH002386 to LEE and R01DK069575 to GGH