Effects of adenoviral overexpression of uncoupling protein-2 and -3 on mitochondrial respiration in insulinoma cells.

The brown adipose tissue uncoupling protein 1 (UCP1) catalyzes proton reentry without ATP synthesis, thereby dissipating energy as heat. In contrast, the function(s) of the recently described homologs, UCP2 and UCP3, are less clear. The aim of the present study was to determine whether overexpressed UCP subtypes affect mitochondrial respiration and substrate oxidation in cultured insulin-secreting INS-1 insulinoma cells. Adenoviral overexpression of UCP2 significantly decreased the ADP/O ratio by 31% and 39% in comparison to beta-galactosidase (beta-gal) or the mitochondrial protein manganese superoxide dismutase (MnSOD), respectively, and increased state 4 respiration in the presence of succinate and oligomycin by 52% and 59% in comparison to beta-gal or MnSOD, respectively. Adenoviral overexpression of UCP3 also decreased the ADP/O ratio by 18% (nonsignificant) and increased state 4 respiration by 24% (nonsignificant) in comparison to ss-gal and significantly decreased the ADP/O ratio by 32% and increased state 4 respiration by 35% in comparison to MnSOD. Both UCP2 and UCP3 expression significantly increased whole cell lipid oxidation by 34% (P < 0.01) and 30% (P < 0.05), respectively, compared with cells expressing Ad5CMVlacZ. However, glucose oxidation was not significantly altered by UCP2 or UCP3 expression. Adenoviral UCP2 expression, but not UCP3 (compared with beta-gal), significantly inhibited insulin secretion in the presence of 15 mM glucose [6.17 +/- 0.42 ng/mg cell protein for beta-gal compared with 4.69 +/- 0.39 for UCP2 (P < 0.05) and 5.51 +/- 0.50 for UCP3]. Both overexpressed UCPs significantly reduced INS-1 cell ATP content. Within certain limitations, which are discussed, these data are the first to demonstrate increased respiration and impaired coupling of oxidative phosphorylation as a result of UCP homolog expression in isolated mammalian mitochondria. Our results also suggest an important role for UCP in lipid metabolism and, possibly, insulin secretion.