TRANSCRIPTIONAL ACTIVATION OF THE GLUT2 GENE BY THE IPF-1 STF-1/IDX-1HOMEOBOX FACTOR/

The homeodomain protein PDX-1, referred as IPF-1/STF-1/IDX-1, is a tra nscriptional factor that plays a critical role in the control of sever al genes expressed in the pancreatic islet. PDX-1 gene expression has been previously shown to be reduced in cultured beta-cell lines chroni cally exposed to high glucose concentrations, As the glucose transport er type 2 (GLUT2) gene expression is selectively decreased in the beta -pancreatic cells of experimental models of diabetes, we postulated th at the loss of GLUTS gene expression in the pancreatic islets of diabe tic animals may be due to the loss of PDX-1 transacting function on th e GLUTS gene. We, therefore, investigated the potential role of PDX-1 in the transcriptional control of GLUTS. We have identified a repeat o f a TAAT motif (5'-TAATA-ATAACA-3') conserved in the sequence of the h uman and murine GLUT2 promoters, Recombinant PDX-1 binds to this GLUT2 TAAT motif in electrophoretic mobility shift experiments. PDX-1 antise rum detects the formation of the complex of PDX-1 with the GLUT2TAAT m otif in nuclear extracts from the pancreatic insulin-secreting cell li ne, beta TC3, The GLUT2TAAT motif was mutated in the murine GLUT2 prom oter (-1308/+49 bp) linked to a luciferase reporter gene and transfect ed into beta TC3 cells. Compared with the transcriptional activity of the wild type promoter, that of the mutated promoter decreases by 41%. Multiple copies of the GLUT2TAAT motif were ligated 5' to a heterolog ous promoter and transfected into a PDX-1-expressing cell line (beta T C3) and into cell lines lacking the homeobox factor (InR1-G9 and JEG-3 ). The GLUT2TAAT motif mediates the activation of the heterologous pro moter in the PDX-1-expressing cell line but not in InR1-G9 or JEG-3 ce ll lines, Furthermore, cotransfection in a PDX-1-deficient cell line w ith the expression vector encoding PDX-1 transactivates specifically t he heterologous promoter containing the multimerized GLUT2TAAT motif, These data demonstrate that the murine GLUT2 promoter is controlled by the PDX-1 homeobox factor through the identified GLUT2TAAT motif.