A significant portion of mitochondrial proton leak in intact thymocytes depends on expression of UCP2.
The uncoupling protein homologue UCP2 is expressed in a variety of mammalian cells. It is thought to be an uncoupler of oxidative phosphorylation. Uncoupling proteins previously have been shown to be capable of translocating protons across phospholipid bilayers in proteoliposome systems. Furthermore, studies in mitochondria from yeast overexpressing the proteins have led to suggestions that they may act as uncouplers in cells. However, this issue is controversial, and to date, definitive experimental evidence is lacking as to whether UCP2 mediates part or all of the basal mitochondrial proton leak in mammalian cells in situ. In the present study, by using thymocytes isolated from UCP2-deficient and wild-type (WT) mice, we addressed the question whether UCP2 is directly involved in catalyzing proton leak in intact cells. Over a range of mitochondrial membrane potentials (DeltaPsi(m)), proton leak activity was lower in thymocytes from UCP2-deficient mice compared with WT mice. At physiological levels of DeltaPsi(m), a significant portion (50%) of basal proton leak in resting cells depended on UCP2. Of note, proton leak in whole cells from WT mice, but not UCP2-deficient mice, responded to stimulation by 4-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-napthalenyl)-1-propenyl]benzoic acid (TTNPB), a known activator of UCP2 activity. Consistent with the observed changes in proton leak, DeltaPsi(m) and ATP levels were increased in untreated thymocytes from UCP2-deficient mice. Interestingly, resting respiration was unaltered, suggesting that UCP2 function in resting cells may be concerned with the control of ATP production rather than substrate oxidation. This study establishes that UCP2, expressed at endogenous levels, mediates proton leak in intact cells.
Krauss
Stefan
S
Division of Endocrinology, Department of Medicine, Beth Israel Deaconess Medical Center and Harvard Medical School, 99 Brookline Avenue, Boston, MA 02215, USA.
Zhang
Chen-Yu
CY
Lowell
Bradford B
BB
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D001565:Benzoates / Q000494:pharmacology; D015152:Blotting, Northern; D015704:CD4 Antigens / Q000096:biosynthesis; D016827:CD8 Antigens / Q000096:biosynthesis; D002455:Cell Division; D002469:Cell Separation; D005434:Flow Cytometry; D005838:Genotype; D015151:Immunoblotting; D007202:Indicators and Reagents / Q000494:pharmacology; D007473:Ion Channels; D008564:Membrane Potentials; D026901:Membrane Transport Proteins; D051379:Mice; D018345:Mice, Knockout; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins; D008954:Models, Biological; D009861:Onium Compounds / Q000494:pharmacology; D010100:Oxygen / Q000378:metabolism*; D010101:Oxygen Consumption; D010766:Phosphorylation; D014176:Protein Biosynthesis; D011522:Protons; D012176:Retinoids / Q000494:pharmacology; D013950:Thymus Gland / Q000166:cytology*; D014212:Tretinoin / Q000031:analogs & derivatives; D014320:Triphenylmethyl Compounds / Q000494:pharmacology; D000071246:Uncoupling Protein 2
D000970:Antineoplastic Agents; D001565:Benzoates; D015704:CD4 Antigens; D016827:CD8 Antigens; D007202:Indicators and Reagents; D007473:Ion Channels; D026901:Membrane Transport Proteins; D024101:Mitochondrial Proteins; D009861:Onium Compounds; D011522:Protons; D012176:Retinoids; D014320:Triphenylmethyl Compounds; C000604390:Ucp2 protein, mouse; D000071246:Uncoupling Protein 2; C011561:triphenylmethylphosphonium; D014212:Tretinoin; C038864:4-(2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1-propenyl)benzoic acid; D000255:Adenosine Triphosphate; D010100:Oxygen