Mechanisms for transcriptional activation of Reg (Regenerating gene)

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In 1984, we found that the administration of poly(ADP-ribose) synthetase inhibitors such as nicotinamide (NA) to 90% depancreatized rats induced islet regeneration, thereby ameliorating the surgical diabetes (Diabetes 33, 401). We isolated a gene, Reg (regenerating gene) from the regenerating islet-derived cDNA library (J. Biol. Chem. 263, 2111, 1988) and demonstrated that Reg protein induced β-cell regeneration to ameliorate experimental diabetes (Proc. Natl. Acad. Sci. USA 91, 3589, 1994 ; Endocrinology 139, 2369, 1998). Reg gene is expressed in the regenerative processes of β-cell but not in normal islets. Therefore, the activation of Reg gene is considered to be a key step for β-cell regeneration.In the present study, we found the activation of Reg gene by interleukin (IL)-6, dexamethasone (Dx) and NA in RINm5F β-cells. RT-PCR and immunoblot analyses revealed that the combined addition of IL-6 (5,000 u/ml) and Dx (100 nM) induced Reg gene expression in RINm5F cells, and the inducti … More on was significantly enhanced by the addition of 5 - 10 mM NA. A series of 5'-deletions of rat Reg gene for luciferase assay was transfected into RINm5F cells. Deletion of the sequence from -2303 to -81 caused no significant changes of the induction by IL-6/Dx/NA. Further deletion to -70 caused a marked loss of the induction. Gel shift assay using a series of mutated competitors suggested a GC box-like sequence in the region is involved in the induction. In fact, a two-base mutation construct for luciferase assay in the GC box-like sequence did not show the induction by IL-6/Dx/NA. A 120 kDa protein was revealed to bind the GC box-like sequence by Southwestern analysis, but antibodies against GC box binding proteins such as Sp1-4 failed to show any "super-shift" in the active transcription complex. These results suggest that IL-6 and glucocorticoids produced in regenerative processes such as insulitis activate Reg gene via the binding of the 120 kDa factor to the GC box-like sequence and that β-cell regeneration by NA is achieved by the enhancement of the Reg gene activation. Less