Glucose-dependent translocation of insulin promoter factor-1 (IPF-1) between the nuclear periphery and the nucleoplasm of single MIN6 beta-cells.
Using laser-scanning confocal microscopy, we have monitored glucose-induced changes in the subcellular localization of insulin promoter factor-1 (IPF-1) labeled with a c-myc epitope tag. This construct trans-activated the insulin promoter in single living MIN6-beta-cells as assessed by luciferase-based promoter analysis. IPF-1.c-myc expression also enhanced the response of the insulin promoter to elevations in extracellular glucose concentration. In the majority (148/235, 63%) of cells maintained at low (3 mM) extracellular glucose concentration, IPF-1.c-myc immunoreactivity was confined to the nuclear periphery. Incubation of cells at stimulatory (30 mM) glucose concentrations caused a rapid redistribution of the chimera to the nucleoplasm (775/958, 81% of cells). By contrast, the irrelevant transcription factor c-Fos, tagged with either c-myc or as a chimera with luciferase, was localized exclusively to the nucleoplasm irrespective of the glucose concentration. Furthermore, IPF-1 extended with the bulky (27 kDa) enhanced green fluorescent protein (EGFP) group was confined largely to the nucleoplasm at all glucose concentrations tested and did not support trans-activation of the insulin promoter by glucose. Movement of endogenous IPF-1 from the nuclear periphery to the nucleoplasm may therefore increase the trans-activational capacity of this factor in native beta-cells exposed to high extracellular glucose concentrations.
Rafiq
I
I
Department of Biochemistry, School of Medical Sciences, University Walk, University of Bristol, Bristol BS8 1TD, United Kingdom.
Kennedy
H J
HJ
Rutter
G A
GA
D001692:Biological Transport; D015415:Biomarkers; D002451:Cell Compartmentation; D002467:Cell Nucleus / Q000378:metabolism*; D005947:Glucose / Q000494:pharmacology*; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins; D007091:Image Processing, Computer-Assisted; D007328:Insulin / Q000096:biosynthesis*; D007515:Islets of Langerhans / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
D015415:Biomarkers; D018398:Homeodomain Proteins; D007328:Insulin; D008164:Luminescent Proteins; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein; D049452:Green Fluorescent Proteins; D005947:Glucose