A significant portion of mitochondrial proton leak in intact thymocytes depends on expression of UCP2

Abstract

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 (⌬⌿ m), proton leak activity was lower in thymocytes from UCP2-deficient mice compared with WT mice. At physiological levels of ⌬⌿ 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, ⌬⌿ 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.

Mitochondrial protein mitofusin 2 is required for NLRP3 inflammasome activation after RNA virus infection

“…We next examined whether the suppression of NLRP3 inflammasome activation by chemical treatment can be reproduced by overexpression of uncoupling protein-2 (UCP-2), which is known to reduce ΔΨ(m) by proton leakage (27,28). Transfection of human embryonic kidney cell line 293T (HEK293T) cells with full-length UCP-2 (full) but not a truncated mutant, UCP-2 (1-200), which contains only amino acid residues 1 to 200, diminished ΔΨ(m) in transfected cells ( Fig.…”

Lack of UCP2 reduces fas-mediated liver injury inob/ob mice and reveals importance of cell-specific UCP2 expression

“…[11][12][13] By tapping into the proton gradient, activated UCP2 may compete with ATP synthase for the electrochemical energy of mitochondria and result in altered cellular ATP levels. 14,15 UCP2 expression in healthy liver is primarily localized to Kupffer cells 16 ; however, UCP2 becomes strikingly abundant in hepatocytes of fatty liver. [17][18][19] Although accumulation and peroxidation of lipids in fatty hepatocytes may advance the effects of UCP2, the biological importance of these changes is not well understood.…”

Uncoupling protein-2 deficiency promotes oxidant stress and delays liver regeneration in mice

“…9,10,12,13 Uncoupling protein-2 (UCP2), a recently identified inner mitochondrial membrane anion carrier, 14,15 has emerged as a sensor 16,17 and potentially critical negative regulator of mitochondrial superoxide production. 18,19 UCP2 is able to mediate proton leaks in mammalian cells 20 and keep the membrane potential sufficiently low to minimize superoxide production. 18,19 Loss of UCP2 function results in increased ROS generation as illustrated by various in vitro and in vivo experiments.…”