UCPs — unlikely calcium porters

Abstract

The mitochondrial Ca 2+ uniporter (MCU) has been characterized for several decades [1], but the molecule responsible for this transport activity is currently unknown. Therefore, a recent Nature Cell Biology paper by Trenker et al. [2] reporting a role for uncoupling proteins (UCP2/3) in mitochondrial Ca 2+ uniport generated much excitement in the field. Subsequently, the authors contend that Ca 2+ transport accounts for most physiologic effects assigned to UCPs [3].The defining characteristic of mitochondrial Ca 2+ uptake is that mitochondrial matrix Ca 2+ ([Ca 2+ ] m ) responds to changes in cytosolic Ca 2+ at a rate dependent on both the driving force (the electrochemical gradient comprising membrane potential (Δψ m ) plus Ca 2+ concentration gradient), and the transporter activity [4]. The steady-state free [Ca 2+ ] m depends on the balance between Ca 2+ uptake and efflux [4]. Trenker et al. report that UCP overexpression increased the free [Ca 2+ ] m attained upon agonist stimulation (Fig. 1c in [2]), apparently without effect on the rate of Ca 2+ uptake. When mitochondrial Ca 2+ efflux via the Na + /Ca 2+ exchanger was blocked using CGP37157, and the driving force for Ca 2+ entry (Δψ m ) was collapsed by oligomycin plus FCCP, agonist-stimulated free [Ca 2+ ] m was again higher in UCP overexpressing cells, with no discernible effect on Ca 2+ uptake rate (Fig. 1f in [2]). In such a system, where both Ca 2+ uptake and efflux are inactive, an effect of UCP expression on steady-state free [Ca 2+ ] m levels cannot be due to changes in Ca 2+ transport. Since overexpression of UCPs can sometimes result in a non-functional uncoupled phenotype thought to be due to protein misfolding [5], and the apparent lack of effect of UCP overexpression on Δψ m (supplement Fig. 1d ]), the significance of these data is also difficult to assess.Alternative to the MCU, the Na + /Ca 2+ or H + /Ca 2+ exchangers could serve a mitochondrial Ca 2+ uptake function, in which UCPs may play a role. However, the V max of these systems is 2 or 18 nmol Ca 2+ /min/mg protein for liver or heart mitochondria respectively, i.e. 2-3 orders of magnitude lower than the V max of the MCU (1700 nmol Ca 2+ /min/mg protein) [6,7]. Thus UCPs cannot significantly affect mitochondrial Ca 2+ uptake through such mechanisms.In Fig. 3 of [2], Ca 2+ uptake by isolated murine liver mitochondria was only partially inhibited by 100 nM ruthenium red (RR), although it is not clear whether crude RR was used (IC 50 ~1 µM for MCU activity) or its purified derivative Ru360 (IC 50 5 nM) [4,8]. Using this experimental system, a decrease in Ca 2+ uptake by liver mitochondria of Ucp2 −/− mice was observed relative to wild-type controls. However, liver mitochondria contain exceedingly small amounts of UCP2 (<1 ng UCP2/mg protein, vs. 25 or 75 ng UCP2/mg protein in mouse kidney or spleen mitochondria respectively; V. Azzu and M.D. Brand unpublished observations). Furthermore, the mitochondrial inner membrane surface area is 5 × 10 10 µm 2 / mg protein [9] and each µm 2 cont...

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