Study of the promoter regions of human mitochondria!- Glycerol-3-phosphate dehydrogenase

Mitochondrial glycerol-3-phosphate dehydrogenase (mGPD), the key enzyme of the glycerol-phosphate shuttle, is encoded in the nucleus and is located in the outer surface of the inner mitochondrial membrane. mGPD is abundant in the normal pancreatic islet. The glycerol phosphate shuttle is believed to play a role in the metabolic signaling of insulin secretion in islet beta cells, and the activity of mGPD is low in several rodent models of non-insulin dependent diabetes. Our analyses of the 5'-untranslated region of the human mGPD mRNA and its genomic organization revealed two alternative first exons designated E1A and E1B. Each of these exons is alternatively spliced to a common exon (exon 2). Sequencing of the 5'flanking regions containing the two putative mGPD promoters PE1A and PE1B indicated that they contain no TATA box, but have several binding sites for transcription factors Sp1 and AP-2. We assessed promoter activity in transient expression assays using luciferase as a reporter. Luciferase activity of PE1A was 10% of PE1B in both INS-1 and HeLa cells. Progressive 5' deletions in the region immediately proximal to E1B resulted in sequentially larger decreases in promoter activity. These findings suggest that the human mGPD gene contains at least two promoters that control transcription and provide a molecular basis for the alternative splicing of mGPD transcripts.