title: Mechanisms of action of islet neogenesis-associated protein: comparison of the full-length recombinant protein and a bioactive peptide.
abstract: Islet neogenesis-associated protein (INGAP) was discovered in the partially duct-obstructed hamster pancreas as a factor inducing formation of new duct-associated islets. A bioactive portion of INGAP, INGAP(104-118) peptide (INGAP-P), has been shown to have neogenic and insulin-potentiating activity in numerous studies, including recent phase 2 clinical trials that demonstrated improved glucose homeostasis in both type 1 and type 2 diabetic patients. Aiming to improve INGAP-P efficacy and to understand its mechanism of action, we cloned the full-length protein (rINGAP) and compared the signaling events induced by the protein and the peptide in RIN-m5F cells that respond to INGAP with an increase in proliferation. Here, we show that, although both rINGAP and INGAP-P signal via the Ras/Raf/ERK pathway, rINGAP is at least 100 times more efficient on a molar basis than INGAP-P. For either ligand, ERK1/2 activation appears to be pertussis toxin sensitive, suggesting involvement of a G protein-coupled receptor(s). However, there are clear differences between the peptide and the protein in interactions with the cell surface and in the downstream signaling. We demonstrate that fluorescent-labeled rINGAP is characterized by clustering on the membrane and by slow internalization (≤5 h), whereas INGAP-P does not cluster and is internalized within minutes. Signaling by rINGAP appears to involve Src, in contrast to INGAP-P, which appears to activate Akt in addition to the Ras/Raf/ERK1/2 pathway. Thus our data suggest that interactions of INGAP with the cell surface are important to consider for further development of INGAP as a pharmacotherapy for diabetes.
authors: Department of Surgery, the Research Institute of the McGill University Health Center, McGill University, Montreal, Québec, Canada. maria.petropavlovskaya@mcgill.ca
authors: Maria
authors: M
authors: Petropavlovskaia
authors: Jamal
authors: J
authors: Daoud
authors: Jonathan
authors: J
authors: Zhu
authors: Mandana
authors: M
authors: Moosavi
authors: Jieping
authors: J
authors: Ding
authors: Julia
authors: J
authors: Makhlin
authors: Beatrice
authors: B
authors: Assouline-Thomas
authors: Lawrence
authors: L
authors: Rosenberg
mesh_terms: D000818:Animals; D000951:Antigens, Neoplasm / Q000494:pharmacology*; D014408:Biomarkers, Tumor / Q000494:pharmacology*; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D003001:Cloning, Molecular; D016207:Cytokines / Q000494:pharmacology*; D007515:Islets of Langerhans / Q000187:drug effects*; D037181:Lectins, C-Type; D000073718:Pancreatitis-Associated Proteins; D010446:Peptide Fragments / Q000494:pharmacology*; D037342:Pertussis Toxin / Q000494:pharmacology; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D000951:Antigens, Neoplasm; D014408:Biomarkers, Tumor; D016207:Cytokines; C500438:INGAP peptide; D037181:Lectins, C-Type; D000073718:Pancreatitis-Associated Proteins; D010446:Peptide Fragments; C000612141:REG3A protein, human; D011994:Recombinant Proteins; C000612144:Reg3b protein, rat; D037342:Pertussis Toxin