Glucose-stimulated preproinsulin gene expression and nuclear trans-location of pancreatic duodenum homeobox-1 require activation of phosphatidylinositol 3-kinase but not p38 MAPK/SAPK2.
Exposure of islet beta-cells to elevated glucose concentrations (30 versus 3 mm) prompts enhanced preproinsulin (PPI) gene transcription and the trans-location to the nucleoplasm of pancreatic duodenum homeobox-1 (PDX-1; Rafiq, I., Kennedy, H., and Rutter, G. A. (1998) J. Biol. Chem. 273, 23241-23247). Here, we show that in MIN6 beta-cells, over-expression of p110.CAAX, a constitutively active form of phosphatidylinositol 3-kinase (PI3K) mimicked the activatory effects of glucose on PPI promoter activity, whereas Deltap85, a dominant negative form of the p85 subunit lacking the p110-binding domain, and the PI3K inhibitor LY 294002, blocked these effects. Similarly, glucose-stimulated nuclear trans-location of endogenous PDX-1 was blocked by Deltap85 expression, and wortmannin or LY 294002 blocked the trans-location from the nuclear membrane to the nucleoplasm of epitope-tagged PDX-1.c-myc. By contrast, SB 203580, an inhibitor of stress-activated protein kinase-2 (SAPK2)/p38 MAP kinase, had no effect on any of the above parameters, and PPI promoter activity and PDX-1.c-myc localization were unaffected by over-expression of the upstream kinase MKK6 (MAP kinase kinase-6) or wild-type p38/SAPK2, respectively. Furthermore, no change in the activity of extracted p38/SAPK2 could be detected after incubation of cells at either 3 or 30 mm glucose. These data suggest that stimulation of PI3K is necessary and sufficient for the effects of glucose on PPI gene transcription, acting via a downstream signaling pathway that does not involve p38/SAPK2.
Rafiq
I
I
Department of Biochemistry, School of Medical Sciences, University Walk, University of Bristol, Bristol BS8 1TD, United Kingdom.
da Silva Xavier
G
G
Hooper
S
S
Rutter
G A
GA
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D002867:Chromones / Q000494:pharmacology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D005947:Glucose / Q000494:pharmacology*; D018398:Homeodomain Proteins / Q000235:genetics*; D007093:Imidazoles / Q000494:pharmacology; D007328:Insulin; D007515:Islets of Langerhans / Q000378:metabolism; D048669:MAP Kinase Kinase 6; D018613:Microscopy, Confocal; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D011384:Proinsulin / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011498:Protein Precursors / Q000235:genetics*; D011725:Pyridines / Q000494:pharmacology; D015398:Signal Transduction; D015534:Trans-Activators / Q000235:genetics*; D014162:Transfection; D000077191:Wortmannin; D048051:p38 Mitogen-Activated Protein Kinases
D000730:Androstadienes; D002867:Chromones; D004791:Enzyme Inhibitors; D018398:Homeodomain Proteins; D007093:Imidazoles; D007328:Insulin; D009025:Morpholines; D011498:Protein Precursors; D011725:Pyridines; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C013737:preproinsulin; D011384:Proinsulin; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048669:MAP Kinase Kinase 6; D005947:Glucose; C093642:SB 203580; D000077191:Wortmannin