title: Prolactin modulates the association and phosphorylation of SNARE and kinesin/MAP-2 proteins in neonatal pancreatic rat islets.
abstract: Prolactin induces maturation of insulin secretion in cultured neonatal rat islets. In this study, we investigated whether the improved secretory response to glucose caused by prolactin involves alteration in the expression, association and phosphorylation of several proteins that participate in these processes. Messenger RNA was extracted from neonatal rat islets cultured for 5 days in the presence of prolactin and reverse transcribed. Gene expression was analyzed by semi-quantitative RT-PCR and by Western blotting for proteins. The gene transcription and protein expression of kinesin and MAP-2 were increased in prolactin-treated islets compared to the controls. The association and phosphorylation of proteins was analyzed by immunoprecipitation followed by Western blotting, after acute exposure to prolactin. Prolactin increased the association between SNARE proteins and kinesin/MAP-2 while the association of munc-18/syntaxin 1A was decreased. Serine phosphorylation of SNAP-25, syntaxin 1A, munc-18, MAP-2 was significantly higher whereas kinesin phosphorylation was decreased in prolactin-treated islets. There was an increase in SNARE complex formation in islets stimulated with prolactin, 22 mM glucose, 40 mM K(+), 200 microM carbachol and 1 microM PMA. The prolactin-induced increase in the formation of SNARE complex and syntaxin 1A phosphorylation was inhibited by PD098059 and U0126, inhibitors of the MAPK pathway. These findings indicate that prolactin primes pancreatic beta-cells to release insulin by increasing the expression and phosphorylation/association of proteins implicated in the secretory machinery and the MAPK/PKC pathway is important for this effect.
authors.affiliation: Department of Physiology and Biophysics, Institute of Biology, State University of Campinas (UNICAMP), CP 6109, Campinas 13083-970, SP, Brazil.
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Cunha
authors.forename: Letícia P
authors.initials: LP
authors.lastname: Roma
authors.forename: Antonio C
authors.initials: AC
authors.lastname: Boschero
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism*; D016547:Kinesins / Q000235:genetics / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D051938:Munc18 Proteins / Q000235:genetics / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D011388:Prolactin / Q000494:pharmacology*; D011493:Protein Kinase C / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D050600:SNARE Proteins / Q000378:metabolism*; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism; D050827:Syntaxin 1 / Q000378:metabolism; D013997:Time Factors
chemical_list: D007328:Insulin; C489787:MAP2 protein, rat; D008869:Microtubule-Associated Proteins; D051938:Munc18 Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D050600:SNARE Proteins; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D011388:Prolactin; D011493:Protein Kinase C; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D016547:Kinesins; D005947:Glucose