Biochemical Foundations of Health and Energy Conservation in Hibernating Free-ranging Subadult Brown Bear Ursus arctos

Abstract

Brown bears (Ursus arctos) hibernate for 5-7 months without eating, drinking, urinating, and defecating at a metabolic rate of only 25% of the summer activity rate. Nonetheless, they emerge healthy and alert in spring. We quantified the biochemical adaptations for hibernation by comparing the proteome, metabolome, and hematological features of blood from hibernating and active free-ranging subadult brown bears with a focus on conservation of health and energy. We found that total plasma protein concentration increased during hibernation, even though the concentrations of most individual plasma proteins decreased, as did the white blood cell types. Strikingly, antimicrobial defense proteins increased in concentration. Central functions in hibernation involving the coagulation response and protease inhibition, as well as lipid transport and metabolism, were upheld by increased levels of very few key or broad specificity proteins. The changes in coagulation factor levels matched the changes in activity measurements. A dramatic 45-fold increase in sex hormone-binding globulin levels during hibernation draws, for the first time, attention to its significant but unknown role in maintaining hibernation physiology. We propose that energy for the costly protein synthesis is reduced by three mechanisms as follows: (i) dehydration, which increases protein concentration without de novo synthesis; (ii) reduced protein degradation rates due to a 6 °C reduction in body temperature and decreased protease activity; and (iii) a marked redistribution of energy resources only increasing de novo synthesis of a few key proteins. The comprehensive global data identified novel biochemical strategies for bear adaptations to the extreme condition of hibernation and have implications for our understanding of physiology in general.

Keywords: antimicrobial proteins; blood constituents; coagulation factor; complement system; hibernation physiology; metabolomics; protein turnover; proteomics; sex hormone-binding globulin (SHBG).

FIGURE 1.

Protein and metabolite changes in blood plasma in free-ranging brown bears during hibernation in winter and in the same bears when active in summer. A, absorption spectra of undiluted plasma samples collected in winter (W) and summer (S) from seven brown bears (bear IDs at top right. M, male; F, female). All were 3-year-old, except for the 2-year-old 0910M. Top, total protein at 280 nm; bottom, hemoglobin at 415 nm (supplemental Table S5). B, principal component analyses of plasma proteins, amines, and phospholipids showed a clear separation of winter (red) and summer (blue) plasma. C, volcano plots of W/S ratios of plasma proteins, amines, and phospholipids versus p values from paired t tests. Dashed lines at p = 0.05. D, immunoblot of SHBG confirming the large increase in hibernation (proteomics detected 45-fold increased levels in winter plasma (p = 0.0006)). The arrows indicate two forms of glycosylated SHBG (like in human SHBG (28)). Heavy and light chains of bear IgG were ∼52 and 27 kDa. All seven bears analyzed in 2010 showed identical results, which were confirmed by samples collected from subadult bears in 2012 and 2013.

FIGURE 2.

Interplay and regulation of plasma protein factors in coagulation, fibrinolysis, and complement defense in hibernating brown bears. Protein names come from the corresponding human genes. The mean of winter/summer (W/S) ratios of protein levels for each of seven subadult bears and the p values from paired t tests quantified by mass spectrometry have been summarized and illustrated by squared frames. Thick frame, p value < 0.05. W/S ratio increased to >1.2-fold in hibernation (red); moderate W/S ratio changes 0.8–1.2-fold (black); W/S ratio decreased to < 0.8-fold in hibernation (blue). Component not detected (dashed). Component absent in the bear protein database (yellow). Coagulation factors quantified by functional assays are shown in space filling color, red increased and blue decreased in hibernation (Table 1). The figure was modified after Reactome's human pathways (54).