Induction of Endogenous Uncoupling Protein 3 Suppresses Mitochondrial Oxidant Emission during Fatty Acid-supported Respiration

Ethan J Anderson,Hanae Yamazaki,P Darrell Neufer

doi:10.1074/jbc.m706129200

Abstract

Uncoupling protein 3 (UCP3) expression increases dramatically in skeletal muscle under metabolic states associated with elevated lipid metabolism, yet the function of UCP3 in a physiological context remains controversial. Here, in situ mitochondrial H(2)O(2) emission and respiration were measured in permeabilized fiber bundles prepared from both rat and mouse (wild-type) gastrocnemius muscle after a single bout of exercise plus 18 h of recovery (Ex/R) that induced a approximately 2-4-fold increase in UCP3 protein. Elevated uncoupling activity (i.e. GDP inhibitable) was evident in Ex/R fibers only upon the addition of palmitate (known activator of UCP3) or under substrate conditions eliciting substantial rates of H(2)O(2) production (i.e. respiration supported by succinate or palmitoyl-L-carnitine/malate but not pyruvate/malate), indicative of UCP3 activation by endogenous reactive oxygen species. In mice completely lacking UCP3 (ucp3(-/-)), Ex/R failed to induce uncoupling activity. Surprisingly, when UCP3 activity was inhibited by GDP (rats) or in the absence of UCP3 (ucp3(-/-)), H(2)O(2) emission was significantly (p < 0.05) higher in Ex/R versus non-exercised control fibers. Collectively, these findings demonstrate that the oxidant emitting potential of mitochondria is increased in skeletal muscle during recovery from exercise, possibly as a consequence of prolonged reliance on lipid metabolism and/or altered mitochondrial biochemistry/morphology and that induction of UCP3 in vivo mediates an increase in uncoupling activity that restores mitochondrial H(2)O(2) emission to non-exercised, control levels.

Highlights

Conductance across the mitochondrial inner membrane, effectively accelerating and uncoupling respiration from ATP production to produce heat [4, 5]
Skeletal Muscle Uncoupling protein 3 (UCP3) mRNA and Protein Expression during Recovery from Exercise—UCP3 mRNA increased (p Ͻ 0.05) in response to exercise, peaking 4 h after exercise at ϳ8-fold over control in RG and ϳ20-fold over control in WG (Fig. 2A). In both RG and WG, UCP3 mRNA remained significantly elevated through 18 h of recovery but returned to near control levels after 24 h of recovery
The results from the present study demonstrate that a physiological increase in endogenous UCP3 expression induced by acute exercise/recovery attenuates mitochondrial H2O2 emission and accelerates state 4 respiration in skeletal muscle of both rats and mice

Summary

Introduction

Conductance across the mitochondrial inner membrane, effectively accelerating and uncoupling respiration from ATP production to produce heat [4, 5]. An increase in the reliance of skeletal muscle on lipid metabolism is common to each of these metabolic conditions and has led to the hypothesis that UCP3 may function to protect against lipotoxicity in the mitochondrial matrix by catalyzing the efflux of surplus non-activated fatty acids (i.e. lacking CoA) and/or potentially damaging fatty acid peroxides (16 – 20) In this model net proton influx (i.e. uncoupling) associated with the cycling of protonated/deprotonated fatty acids is viewed as a consequence rather than an implicit function of UCP3. Our findings reveal that skeletal myofibers during recovery from exercise display an overall increase in mitochondrial oxidantemitting potential under basal state 4 conditions This increase in the potential for free radical generation is offset by an increase in the expression of UCP3, which when activated by fatty acid and/or mitochondrial ROS, mediates an increase in uncoupling activity that suppresses H2O2 emission to levels comparable with controls

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Results

Conclusion