Abstract
The mitochondrial uncoupling protein‐3 is a member of the mitochondrial carrier protein family. As a homologue of the thermogenic brown fat uncoupling protein‐1, it possesses a mitochondrial uncoupling activity and thus can influence cell energy metabolism but its exact biological function remains unclear. In the present study, uncoupling protein‐3 was expressed in 293 cells using the tetracycline‐inducible system and its impact on cell bioenergetics and responsiveness to the apoptotic stimulus was determined. The induction of uncoupling protein‐3 expression in mitochondria did not lead to uncontrolled respiratory uncoupling in intact cells. However, it caused a GDP‐inhibition of state 4 respiration and a GDP‐induced re‐polarization of the inner mitochondrial membrane in the presence of fatty acids, in agreement with its expected physiological behavior as an uncoupling protein (UCP). Uncoupling protein‐3 expression did not cause apoptosis per se but increased the responsiveness of the cells to a mitochondrial apoptotic stimulus (i.e., addition of staurosporine in the culture medium). It enhanced caspase 3 and caspase 9 activation and favored cytochrome c release. Moreover, cells in which uncoupling protein‐3 expression had been induced showed a higher mitochondrial Bax/Bcl‐2 ratio essentially due to enhanced translocation of Bax from cytosol to mitochondria. Finally, the induction of uncoupling protein‐3 also increased the sensitivity of mitochondria to open the permeability transition pore in response to calcium. It is concluded that the presence of uncoupling protein‐3 in mitochondria sensitizes cells to apoptotic stimuli involving mitochondrial pathways. © 2004 Wiley‐Liss, Inc.