Functional analysis of two promoters for the human mitochondrial glycerol phosphate dehydrogenase gene

Mitochondrial glycerol phosphate dehydrogenase (mGPD) is abundant in the no rmal pancreatic insulin cell, but its level is lowered 50% by diabetes. To evaluate mGPD expression, we cloned and characterized the 5'-flanking regio n of the human mGPD gene. The gene has two alternative first exons and two promoters. The downstream promoter (B) is 10 times more active than the ups tream promoter (A) in insulin secreting cells (INS-1) and HeLa cells. Promo ter B has higher activity in INS-1 than in non-beta cells. Deletion and mut ation analysis suggested that a NRF-2 binding site at -94 to -101 and an E2 F binding site at -208 to -215 are important regulatory cis elements in pro moter B, Gel mobility shift assays indicated that the -94 to -101 region bi nds the NRF-2 protein. When INS-1 cells were maintained in the presence of high glucose (25 mM) for 7 days, mGPD was the only 1 of 6 enzyme activities lowered (53%), mGPD promoter B activity was reduced by 60% in INS-1 cells by the high glucose, but in HepG2 cells and HeLa cells, promoter B activity was unchanged or slightly increased. Deletion analysis indicated the gluco se responsiveness was distributed across the region from -340 to -260 in pr omoter B. The results indicate that mGPD gene transcription in the beta cel l is regulated differently from other cells and that decreased mGPD promote r B transcription is at least in part the cause of the decreased beta cell mGPD levels in diabetes.