Importin beta1 mediates the glucose-stimulated nuclear import of pancreatic and duodenal homeobox-1 in pancreatic islet beta-cells (MIN6).
The transcription factor PDX-1 (pancreatic and duodenal homeobox-1) is essential for pancreatic development and the maintainence of expression of islet beta-cell-specific genes. In an previous study [Rafiq, Kennedy and Rutter (1998) J. Biol. Chem. 273, 23241-23247] we demonstrated that PDX-1 may be activated at elevated glucose concentrations by translocation from undefined binding sites in the cytosol and nuclear membrane into the nucleoplasm. In the present study, we show that PDX-1 interacts directly and specifically in vitro with the nuclear import receptor family member, importin beta1, and that this interaction is mediated by the PDX-1 homeodomain (amino acids 146-206). Demonstrating the functional importance of the PDX-1-importin beta1 interaction, microinjection of MIN6 beta-cells with anti-(importin beta1) antibodies blocked both the nuclear translocation of PDX-1, and the activation by glucose (30 mM versus 3 mM) of the pre-proinsulin promoter. However, treatment with extracts from pancreatic islets incubated at either low or high glucose concentrations had no impact on the ability of PDX-1 to interact with importin beta1 in vitro. Furthermore, importin beta1 also interacted with SREBP1c (sterol-regulatory-element-binding protein 1c) in vitro, and microinjection of importin beta1 antibodies blocked the activation by glucose of SREBP1c target genes. Since the subcellular distribution of SREBP1c is unaffected by glucose, these findings suggest that a redistribution of importin beta1 is unlikely to explain the glucose-stimulated nuclear uptake of PDX-1. Instead, we conclude that the uptake of PDX-1 into the nucleoplasm, as glucose concentrations increase, may be mediated by release of the factor both from sites of retention in the cytosol and from non-productive complexes with importin beta1 at the nuclear membrane.
Guillemain
Ghislaine
G
Henry Wellcome Laboratories for Integrated Cell Signalling and Department of Biochemistry, University of Bristol, Bristol BS8 1TD, UK.
Da Silva Xavier
Gabriela
G
Rafiq
Imran
I
Leturque
Armelle
A
Rutter
Guy A
GA
D021581:Active Transport, Cell Nucleus; D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology*; D018398:Homeodomain Proteins; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D017434:Protein Structure, Tertiary; D051381:Rats; D017208:Rats, Wistar; D051780:Sterol Regulatory Element Binding Protein 1; D015534:Trans-Activators / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors; D014158:Transcription, Genetic; D028961:beta Karyopherins / Q000502:physiology*
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C424166:KPNB1 protein, human; C497389:Srebf1 protein, rat; D051780:Sterol Regulatory Element Binding Protein 1; D015534:Trans-Activators; D014157:Transcription Factors; D028961:beta Karyopherins; C084010:pancreatic and duodenal homeobox 1 protein; D005947:Glucose