UCP2, UCP3, avUCP, what do they do when proton transport is not stimulated? Possible relevance to pyruvate and glutamine metabolism

Uncoupling proteins (UCPs) are specialized members of the mitochondrial transporter family. They allow passive proton transport through the mitochondrial inner membrane. This activity leads to uncoupling of mitochondrial respiration and to energy waste, which is well documented with UCP1 in brown adipose tissue. The uncoupling activity of the new UCPs (discovered after 1997), such as UCP2 and UCP3 in mammals or avUCP in birds, is more difficult to characterize. However, extensive data support the idea that the new UCPs are involved in the control of reactive oxygen species (ROS) generation. This fits with the hypothesis that mild uncoupling caused by the UCPs prevents ROS production. Activators and inhibitors regulate the proton transport activity of the UCPs. In the absence of activators of proton transport, the UCP allows the permeation of other ions. We suggest that this activity has physiological significance and, for example, UCP3 expressed in glycolytic muscle fibres may be a passive pyruvate transporter ensuring equilibrium between glycolysis and oxidative phosphorylation. Induction of UCP2 expression by glutamine strengthens the proposal that new UCPs could act to determine the choice of mitochondrial substrate. This would obviously have an impact on mitochondrial bioenergetics and ROS production.     

Canine mitochondrial uncoupling proteins: structure and mRNA expression of three isoforms in adult beagles

Uncoupling proteins (UCPs) are members of the mitochondrial transporter family that dissipate the proton gradient as heat more than via ATP synthesis. In the present study, nucleotide and amino acid sequences of UCPs 1, 2 and 3 of a dog were determined, and their mRNA expression in various peripheral tissues was examined. The sequences were highly (76-97%) homologous to those of other species. Although lower homologies (60-74%) were found when compared among the three canine UCPs, their deduced amino acid sequences had some common domains, such as three mitochondrial carrier protein motifs, six transmembrane alpha-helix domains, and putative purine nucleotide binding domains. By Northern blot analyses, UCP1 mRNA was not detected in any tissues examined. UCP2 mRNA was expressed in most tissues, particularly abundantly in adipose tissue, spleen and lung. Two sizes of UCP3 mRNA were found exclusively in heart and skeletal muscle. These results suggest that canine UCPs have uncoupling activity, and are involved in the regulation of metabolic heat production and/or energy expenditure, as do those of other species.     

The role of uncoupling proteins in pathophysiological states

Until very recently, the uncoupling protein-1 (UCP1), present only in brown adipose tissue (BAT), was considered to be the only mitochondrial carrier protein that stimulated heat production by dissipating the proton gradient generated during respiration across the inner mitochondrial membrane and therefore uncoupling respiration from ATP synthesis. Recently, new uncoupling proteins, UCP2, UCP3, and UCP4, and brain mitochondrial carrier protein-1 (BMCP-1) have been described in mammalian tissues. The present review deals with the possible role of these proteins in different pathological conditions involving alterations in energy balance such as obesity or cachexia. In conclusion, the emergence of the UCP family has altered the approaches to bioenergetics and stressed the importance of uncoupling respiration in different pathophysiological conditions. An extensive qualitative and quantitative characterization of the new members of the UCP family in mammalian tissues will allow a better understanding of the molecular and regulatory mechanisms of thermogenesis and energy metabolism. At this point, we hope that the knowledge presented in the present review will not only stimulate a debate about the role of the UCP family in disease but also lead to applications beneficial for human health.     

Mitochondrial Uncoupling Proteins in Mammals and Plants

Uncoupling proteins (UCPs) belong to a distinct cluster of the mitochondrial anion carrier family. Up to five different uncoupling protein types were found in mitochondria of mammals and plants, and recently in fishes, fungi and protozoa. They exhibit a significantly conserved structure with several motifs specific to either the whole cluster or protein type. Uncoupling proteins, as well as the whole mitochondrial anion carrier gene family, probably emerged in evolution before the separation of animal, fungi, and plant kingdoms and originate from an anion/nucleotide or anion/anion transporter ancestor. Mammalian UCP1, UCP2, UCP3, and plant uncoupling proteins pUCP1 and pUCP2 are similar and seem to form one subgroup, whereas UCP4 and BMCP1 belong to a different group. Molecular, biochemical, and phylogenic data suggest that UCP2 could be considered as an UCP-prototype. UCP1 plays its biological role mainly in the non-shivering thermogenesis while the role of the other types is unknown. However, hypotheses have suggested that they are involved in the general balance of basic energy expenditure, protection from reactive oxygen species, and, in plants, in fruit ripening and seed ontogeny.     

Respiratory uncoupling by UCP1 and UCP2 and superoxide generation in endothelial cell mitochondria

Mitochondria represent a major source of reactive oxygen species (ROS), particularly during resting or state 4 respiration wherein ATP is not generated. One proposed role for respiratory mitochondrial uncoupling proteins (UCPs) is to decrease mitochondrial membrane potential and thereby protect cells from damage due to ROS. This work was designed to examine superoxide production during state 4 (no ATP production) and state 3 (active ATP synthesis) respiration and to determine whether uncoupling reduced the specific production of this radical species, whether this occurred in endothelial mitochondria per se, and whether this could be modulated by UCPs. Superoxide formation by isolated bovine aortic endothelial cell (BAE) mitochondria, determined using electron paramagnetic resonance spectroscopy, was approximately fourfold greater during state 4 compared with state 3 respiration. UCP1 and UCP2 overexpression both increased the proton conductance of endothelial cell mitochondria, as rigorously determined by the kinetic relationship of respiration to inner membrane potential. However, despite uncoupling, neither UCP1 nor UCP2 altered superoxide formation. Antimycin, known to increase mitochondrial superoxide, was studied as a positive control and markedly enhanced the superoxide spin adduct in our mitochondrial preparations, whereas the signal was markedly impaired by the powerful chemical uncoupler p-(trifluoromethoxyl)-phenyl-hydrazone. In summary, we show that UCPs do have uncoupling properties when expressed in BAE mitochondria but that uncoupling by UCP1 or UCP2 does not prevent acute substrate-driven endothelial cell superoxide as effluxed from mitochondria respiring in vitro.     

Further support to the uncoupling-to-survive theory: the genetic variation of human UCP genes is associated with longevity

In humans Uncoupling Proteins (UCPs) are a group of five mitochondrial inner membrane transporters with variable tissue expression, which seem to function as regulators of energy homeostasis and antioxidants. In particular, these proteins uncouple respiration from ATP production, allowing stored energy to be released as heat. Data from experimental models have previously suggested that UCPs may play an important role on aging rate and lifespan. We analyzed the genetic variability of human UCPs in cohorts of subjects ranging between 64 and 105 years of age (for a total of 598 subjects), to determine whether specific UCP variability affects human longevity. Indeed, we found that the genetic variability of UCP2, UCP3 and UCP4 do affect the individual's chances of surviving up to a very old age. This confirms the importance of energy storage, energy use and modulation of ROS production in the aging process. In addition, given the different localization of these UCPs (UCP2 is expressed in various tissues including brain, hearth and adipose tissue, while UCP3 is expressed in muscles and Brown Adipose Tissue and UCP4 is expressed in neuronal cells), our results may suggest that the uncoupling process plays an important role in modulating aging especially in muscular and nervous tissues, which are indeed very responsive to metabolic alterations and are very important in estimating health status and survival in the elderly.     

Mitochondrial uncoupling proteins in the CNS: in support of function and survival

Mitochondrial uncoupling mediated by uncoupling protein 1 (UCP1) is classically associated with non-shivering thermogenesis by brown fat. Recent evidence indicates that UCP family proteins are also present in selected neurons. Unlike UCP1, these proteins (UCP2, UCP4 and BMCP1/UCP5) are not constitutive uncouplers and are not crucial for non-shivering thermogenesis. However, they can be activated by free radicals and free fatty acids, and their activity has a profound influence on neuronal function. By regulating mitochondrial biogenesis, calcium flux, free radical production and local temperature, neuronal UCPs can directly influence neurotransmission, synaptic plasticity and neurodegenerative processes. Insights into the regulation and function of these proteins offer unsuspected avenues for a better understanding of synaptic transmission and neurodegeneration.     

Control of Mitochondrial pH by Uncoupling Protein 4 in Astrocytes Promotes Neuronal Survival

Brain activity is energetically costly and requires a steady and highly regulated flow of energy equivalents between neural cells. It is believed that a substantial share of cerebral glucose, the major source of energy of the brain, will preferentially be metabolized in astrocytes via aerobic glycolysis. The aim of this study was to evaluate whether uncoupling proteins (UCPs), located in the inner membrane of mitochondria, play a role in setting up the metabolic response pattern of astrocytes. UCPs are believed to mediate the transmembrane transfer of protons, resulting in the uncoupling of oxidative phosphorylation from ATP production. UCPs are therefore potentially important regulators of energy fluxes. The main UCP isoforms expressed in the brain are UCP2, UCP4, and UCP5. We examined in particular the role of UCP4 in neuron-astrocyte metabolic coupling and measured a range of functional metabolic parameters including mitochondrial electrical potential and pH, reactive oxygen species production, NAD/NADH ratio, ATP/ADP ratio, CO₂ and lactate production, and oxygen consumption rate. In brief, we found that UCP4 regulates the intramitochondrial pH of astrocytes, which acidifies as a consequence of glutamate uptake, with the main consequence of reducing efficiency of mitochondrial ATP production. The diminished ATP production is effectively compensated by enhancement of glycolysis. This nonoxidative production of energy is not associated with deleterious H₂O₂ production. We show that astrocytes expressing more UCP4 produced more lactate, which is used as an energy source by neurons, and had the ability to enhance neuronal survival.     

4-hydroxy-2-nonenal and uncoupling proteins: an approach for regulation of mitochondrial ROS production

One factor that has the potential to regulate reactive oxygen species (ROS) generation is the mild uncoupling of oxidative phosphorylation, i.e. proton (H(+)) leak across the mitochondrial inner membrane. Proton leak has been shown to attenuate ROS generation, whereas ROS and their derivatives (such as superoxide and hydroxynonenal) have been shown to induce H(+) leak through uncoupling proteins (UCPs). This suggests the existence of a feedback loop between ROS and H(+) leak mediated through UCPs. Although the physiological functions of the new UCPs, such as UCP2 and UCP3, are still not established, extensive data support the idea that these mitochondrial carrier proteins are involved in the control of ROS generation. The molecular basis of both ROS generation and hydroxynonenal-induced uncoupling through UCPs is reviewed and the consequences of their interaction for protection against excessive ROS production at the expense of energy production is discussed.     

Respiration under control of uncoupling proteins: Clinical perspective

The term 'uncoupling protein' was originally used for the mitochondrial membrane protein UCP1, which is uniquely present in mitochondria of brown adipocytes, thermogenic cells that regulate body temperature in small rodents, hibernators and mammalian newborns. In these cells, UCP1 acts as a proton carrier activated by free fatty acids and creates a shunt between complexes of the respiratory chain and ATP-synthase resulting in a futile proton cycling and dissipation of oxidation energy as heat. Recent identification of new homologues to UCP1 expressed in brown and white adipose tissue, muscle, brain and other tissues together with the hypothesis that these novel uncoupling proteins (UCPs) may regulate thermogenesis and/or fatty acid metabolism and furthermore may protect against free radical oxygen species production have generated considerable optimism for rapid advances in the identification of new targets for pharmacological management of complex pathological syndromes such as obesity, type 2 diabetes or chronic inflammatory diseases. However, since the physiological and biochemical roles of the novel UCPs are not yet clear, the main challenge today consists first of all in providing mechanistic explanation for their functions in cellular physiology. This lively awaited information may be the basis for potential pharmacological targeting of the UCPs in future.