In response to hypoxic stress, many animals compensate for a reduced cellular O 2 supply by suppressing total metabolism, thereby reducing O 2 demand. For small endotherms that are native to high-altitude environments, this is not always a viable strategy, as the capacity for sustained aerobic thermogenesis is critical for survival during periods of prolonged cold stress. For example, survivorship studies of deer mice (Peromyscus maniculatus) have demonstrated that thermogenic capacity is under strong directional selection at high altitude. Here, we integrate measures of whole-organism thermogenic performance with measures of metabolic enzyme activities and genomic transcriptional profiles to examine the mechanistic underpinnings of adaptive variation in this complex trait in deer mice that are native to different elevations. We demonstrate that highland deer mice have an enhanced thermogenic capacity under hypoxia compared with lowland conspecifics and a closely related lowland species, Peromyscus leucopus. Our findings suggest that the enhanced thermogenic performance of highland deer mice is largely attributable to an increased capacity to oxidize lipids as a primary metabolic fuel source. This enhanced capacity for aerobic thermogenesis is associated with elevated activities of muscle metabolic enzymes that influence flux through fatty-acid oxidation and oxidative phosphorylation pathways in high-altitude deer mice and by concomitant changes in the expression of genes in these same pathways. Contrary to predictions derived from studies of humans at high altitude, our results suggest that selection to sustain prolonged thermogenesis under hypoxia promotes a shift in metabolic fuel use in favor of lipids over carbohydrates.functional genomics | RNA-seq | thermoregulation | transcriptomics D uring cold stress, homeothermic endotherms maintain a constant body temperature by increasing metabolic heat production. In small endotherms like mice that have high thermoregulatory demands, thermogenic capacity influences survival in cold environments and therefore has a clear connection to Darwinian fitness (1, 2). Indeed, thermogenic capacity in freeranging deer mice (Peromyscus maniculatus) is subject to strong directional selection at high altitude (3).Sustaining maximal thermogenic capacities during prolonged periods of cold stress requires a high rate of O 2 flux through oxidative pathways, and this requirement presents a unique challenge for endothermic animals that live under conditions of chronic O 2 deprivation at high altitude. The reduced partial pressure of O 2 (PO 2 ) at high altitude imposes well-documented constraints on aerobic metabolism (4-8), thereby exacerbating the increased thermoregulatory demands faced by endothermic animals that are native to cold, alpine environments.In rodents, aerobic thermogenesis is accomplished through both shivering and nonshivering mechanisms, and in deer mice, shivering accounts for roughly 35-50% of total thermogenic capacity (9). As with other forms of strenuous exerci...

SUMMARYSmall mammals face especially severe thermoregulatory challenges at high altitude because the reduced O 2 availability constrains the capacity for aerobic thermogenesis. Adaptive enhancement of thermogenic performance under hypoxic conditions may be achieved via physiological adjustments that occur within the lifetime of individuals (phenotypic plasticity) and/or genetically based changes that occur across generations, but their relative contributions to performance differences between highland and lowland natives are unclear. Here, we examined potentially evolved differences in thermogenic performance between populations of deer mice (Peromyscus maniculatus) that are native to different altitudes. The purpose of the study was to assess the contribution of phenotypic plasticity to population differences in thermogenic performance under hypoxia. We used a common-garden deacclimation experiment to demonstrate that highland deer mice have enhanced thermogenic capacities under hypoxia, and that performance differences between highland and lowland mice persist when individuals are born and reared under common-garden conditions, suggesting that differences in thermogenic capacity have a genetic basis. Conversely, population differences in thermogenic endurance appear to be entirely attributable to physiological plasticity during adulthood. These combined results reveal distinct sources of phenotypic plasticity for different aspects of thermogenic performance, and suggest that thermogenic capacity and endurance may have different mechanistic underpinnings. Supplementary material available online at

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A mechanistic understanding of the response of metabolic rate to temperature is essential for understanding thermal ecology and metabolic adaptation. Although the Arrhenius equation has been used to describe the effects of temperature on reaction rates and metabolic traits, it does not adequately describe two aspects of the thermal performance curve (TPC) for metabolic rate—that metabolic rate is a unimodal function of temperature often with maximal values in the biologically relevant temperature range and that activation energies are temperature dependent. We show that the temperature dependence of metabolic rate in ectotherms is well described by an enzyme‐assisted Arrhenius (EAAR) model that accounts for the temperature‐dependent contribution of enzymes to decreasing the activation energy required for reactions to occur. The model is mechanistically derived using the thermodynamic rules that govern protein stability. We contrast our model with other unimodal functions that also can be used to describe the temperature dependence of metabolic rate to show how the EAAR model provides an important advance over previous work. We fit the EAAR model to metabolic rate data for a variety of taxa to demonstrate the model's utility in describing metabolic rate TPCs while revealing significant differences in thermodynamic properties across species and acclimation temperatures. Our model advances our ability to understand the metabolic and ecological consequences of increases in the mean and variance of temperature associated with global climate change. In addition, the model suggests avenues by which organisms can acclimate and adapt to changing thermal environments. Furthermore, the parameters in the EAAR model generate links between organismal level performance and underlying molecular processes that can be tested for in future work.

The repeatability of a trait is a critical factor in determining how that trait is affected by natural selection. We examined the repeatability of a key physiological trait, maximum oxygen consumption (O 2max), in a wild population of Belding's Ground Squirrels, Spermophilus beldingi. O 2max is an integrated measure of organismal metabolic performance. It can be intuitively related to fitness because it sets an upper limit to sustainable power output during ecologically important activites such as locomotion and thermoregulatory heat production. 2. We used respirometry to determine O 2max during exercise and thermogenesis. Exercise O 2max was elicited in an enclosed running wheel. Thermogenic O 2max was obtained with acute cold exposure in a helium-oxygen gas mixture. 3. Repeatability of both exercise and thermogenic O 2max was high over 2 h intervals but declined over longer test periods (6-18 days and 1-2 years). In general, repeatability was higher for exercise O 2max than for thermogenic O 2max. 4. We found no repeatability for animals tested initially as juveniles and then 1 or 2 years later as adults; evidently there is sufficient plasticity in O 2max to decouple aerobic performance between these life stages. A small number of adults tested in successive years showed significant repeatability of exercise O 2max but no repeatability of thermogenic O 2max .