Hobler S C, Williams A, Fischer D, Wang J J, Sun X, Fischer J E, Monaco J J, Hasselgren P O
Department of Surgery, Biochemistry and Microbiology, Howard Hughes Medical Institute, University of Cincinnati, 45267, USA.
Am J Physiol. 1999 Aug;277(2):R434-40. doi: 10.1152/ajpregu.1999.277.2.R434.
Recent studies suggest that sepsis stimulates ubiquitin-dependent protein breakdown in skeletal muscle. In this proteolytic pathway, ubiquitinated proteins are recognized, unfolded, and degraded by the multicatalytic 26S protease complex. The 20S proteasome is the catalytic core of the 26S protease complex. The role of the 20S proteasome in the regulation of sepsis-induced muscle proteolysis is not known. We tested the hypothesis that sepsis increases 20S proteasome activity and the expression of mRNA for various subunits of this complex. Proteolytic activity of isolated 20S proteasomes, assessed as activity against fluorogenic peptide substrates, was increased in extensor digitorum longus muscles from septic rats. The proteolytic activity was inhibited by specific proteasome blockers. Northern blot analysis revealed an approximately twofold increase in the relative abundance of mRNA for the 20S alpha-subunits RC3 and RC9 and the beta-subunit RC7. However, Western blot analysis did not show any difference in RC9 protein content between sham-operated and septic rats. The increased activity and expression of the 20S proteasome in muscles from septic rats lend further support for a role of the ubiquitin-proteasome-pathway in the regulation of sepsis-induced muscle proteolysis.
近期研究表明，脓毒症可刺激骨骼肌中泛素依赖性蛋白质分解。在这条蛋白水解途径中，泛素化蛋白被多催化26S蛋白酶复合体识别、展开并降解。20S蛋白酶体是26S蛋白酶复合体的催化核心。20S蛋白酶体在脓毒症诱导的肌肉蛋白水解调节中的作用尚不清楚。我们检验了脓毒症会增加20S蛋白酶体活性以及该复合体各种亚基的mRNA表达这一假说。通过测定对荧光肽底物的活性来评估，脓毒症大鼠趾长伸肌中分离出的20S蛋白酶体的蛋白水解活性增加。蛋白水解活性受到特异性蛋白酶体阻滞剂的抑制。Northern印迹分析显示，20Sα亚基RC3和RC9以及β亚基RC7的mRNA相对丰度增加了约两倍。然而，蛋白质印迹分析未显示假手术大鼠和脓毒症大鼠之间RC9蛋白含量有任何差异。脓毒症大鼠肌肉中20S蛋白酶体活性和表达的增加，进一步支持了泛素-蛋白酶体途径在脓毒症诱导的肌肉蛋白水解调节中的作用。
