Original Contribution THE CONSEQUENCES OF ACUTE COLD EXPOSURE ON PROTEIN OXIDATION AND PROTEASOME ACTIVITY IN SHORT-TAILED FIELD VOLES, MICROTUS AGRESTIS COLIN SELMAN,* TILMAN GRUNE, † ALEXANDRA STOLZING, † MANUELA JAKSTADT, † JANE S. MCLAREN,* and JOHN R. SPEAKMAN* ‡ *Aberdeen Centre for Energy Expenditure and Obesity (ACERO), Department of Zoology, Aberdeen University, Aberdeen, UK; † Neuroscience Research Centre, Medical Faculty (Charite `), Humboldt-University Berlin, Berlin, Germany; and ‡ ACERO, Division of Appetite and Energy Balance, Rowett Research Institute, Bucksburn, Aberdeen, UK (Received 31 January 2002; Revised 2 April 2002; Accepted 12 April 2002) Abstract—During cold exposure, animals upregulate their metabolism and food intake, potentially exposing them to elevated reactive oxygen species (ROS) production and oxidative damage. We investigated whether acute cold (7  3°C) exposure (1, 10, or 100 h duration) affected protein oxidation and proteasome activity, when compared to warm controls (22  3°C), in a small mammal model, the short-tailed field vole Microtus agrestis. Protein carbonyls and the chymotrypsin-like proteasome activity were measured in plasma, heart, liver, kidney, small intestine (duodenum), skeletal muscle (gastrocnemius), and brown adipose tissue (BAT). Trypsin-like and peptidyl-glutamyl-like proteasome activities were determined in BAT, liver, and skeletal muscle. Resting metabolic rate increased significantly with duration of cold exposure. In skeletal muscle (SM) and liver, protein carbonyl levels also increased with duration of cold exposure, but this pattern was not repeated in BAT where protein carbonyls were not significantly elevated. Chymotrp- sin-like proteasome activity did not differ significantly in any tissue. However, trypsin-like activity in SM and peptidyl-glutamyl-like activity in both skeletal muscle and liver, were reduced during the early phase of cold exposure (1–10 h), correlated with the increased carbonyl levels in these tissues. In contrast there was no reduction in proteasome activity in BAT during the early phase of cold exposure and peptidyl-glutamyl-like activity was significantly increased, correlated with the lack of accumulation of protein carbonyls in this tissue. The upregulation of proteasome activity in BAT may protect this tissue from accumulated oxidative damage to proteins. This protection may be a very important factor in sustaining uncoupled respiration, which underpins nonshivering thermogenesis at cold temperatures. © 2002 Elsevier Science Inc. Keywords—Protein oxidation, Proteasome, Resting metabolic rate, Cold exposure, Free radicals INTRODUCTION Reactive oxygen species (ROS) are produced both as a byproduct of cellular metabolism in aerobic animals and also from various environmental sources [1,2]. When ROS production exceeds the capacities of protection and repair mechanisms oxidative stress occurs, resulting in damage to macromolecules such as proteins, lipids, and DNA [2]. The oxidation of proteins can lead to decline in both protein function and enzyme activity [2–5]. The relative risk of oxidative stress appears to increase during periods of elevated metabolism, including both exercise activity and cold exposure [6 – 8]. In mammalian cells, the breakdown of oxidized pro- teins occurs via two distinct pathways, the lysosomal and proteasomal systems [4,5,9 –11]. The proteasome is a multicatalytic cytosolic and nuclear proteinase complex [12], which appears present in all eukaryotic cells, mi- tochondria, and bacteria [4]. The most widely studied function of the proteasome has been its role in the selective and rapid degradation of oxidized proteins. This rapid degradation, both in the presence and absence of ATP and ubiquitin, appears necessary to reduce the po- tential for aberrant protein accumulation. Indeed, an in- Address correspondence to: Prof. John Speakman, ACERO, Depart- ment of Zoology, University of Aberdeen, Aberdeen, Scotland AB24 2TZ, UK; Tel: +44 (1224) 272879; Fax: +44 (1224) 272396; E-Mail: nhi158@abdn.ac.uk. Free Radical Biology & Medicine, Vol. 33, No. 2, pp. 259 –265, 2002 Copyright © 2002 Elsevier Science Inc. Printed in the USA. All rights reserved 0891-5849/02/$–see front matter PII S0891-5849(02)00874-2 259