VAN BREUKELEN, F.*; QURESHI, S.; Univ. of Nevada, Las Vegas: Depression of ubiquitin-dependent proteolysis during hibernation

Mammalian hibernators cycle between bouts of torpor wherein body temperature may be depressed to below -2° C and euthermic interbout arousals wherein body temperature is approximately 37° C. Protein synthesis is markedly depressed during torpor in concordance with metabolic demands. In the absence of significant protein synthesis, degradation of proteins as part of normal cellular metabolism would rapidly deplete protein pools. Since ubiquitin dependent proteolysis has been implicated in the turnover of most short-lived regulatory proteins, we examined the fate of this system during hibernation. Ubiquitin dependent proteolysis consists of two major steps: 1) the tagging of a protein substrate by ubiquitin and 2) the protein substrate’s subsequent degradation by the proteasome. An earlier study of ubiquitylation during hibernation revealed a 2-3 fold elevation of ubiquitin conjugate concentrations during torpor: an unexpected result that seemingly would suggest increased proteolytic activity. A more likely explanation for these data would be that proteolysis per se was depressed and that the increased levels of ubiquitylated proteins reflect an inability to degrade these tagged proteins. We addressed this question by utilizing an in vitro assay system based on the cleavage of a fluorogenic substrate by the 20S core protease. Preliminary data reveal that the activity of the protease associated with degrading ubiquitin conjugated proteins is extremely temperature sensitive with little to no activity at the temperatures associated with deep torpor. This finding supports the hypothesis that the increased levels of ubiquitylated proteins is explained by a net accumulation due to an inability to degrade the tagged proteins.