Beyette J, Mason G G, Murray R Z, Cohen G M, Rivett A J
Department of Biochemistry, University of Leicester, Leicester LE1 7RH, U.K. and Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK.
Biochem J. 1998 Jun 1;332 ( Pt 2)(Pt 2):315-20. doi: 10.1042/bj3320315.
The induction of apoptosis in thymocytes by the glucocorticoid dexamethasone was used as a model system to investigate whether there are changes in 20 S and 26 S proteasome activities during apoptosis. We observed that thymocytes contain high concentrations of proteasomes and that following treatment with dexamethasone, cell extracts showed a decrease in proteasome chymotrypsin-like activity which correlated with the degree of apoptosis observed. The decrease in chymotrypsin-like activity of 20 S and 26S proteasomes was still apparent after these complexes had been partially purified from apoptotic thymocyte extracts and was therefore not due to competition resulting from a general increase in protein turnover. The trypsin-like and peptidylglutamylpeptide hydrolase activities of proteasome complexes were also observed to decrease during apoptosis, but these decreases were reversed by the inhibition of apoptosis by the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone. However, the chymotrypsin-like activity of proteasomes decreased further in the presence of the apoptosis inhibitor. Val-Ala-Asp-fluoromethylketone was found to inhibit the chymotrypsin- and trypsin-like activity of 26 S proteasomes in vitro. The decrease in proteasome activities in apoptosis did not appear to be due to a decrease in the concentration of total cellular proteasomes. Thus, the early decreases in 20 S and 26 S proteasome activities during apoptosis appear to be due to a down-regulation of their proteolytic activities and not to a decrease in their protein concentration. These data suggest that proteasomes may be responsible, in thymocytes, for the turnover of a protein that functions as a positive regulator of apoptosis.
以糖皮质激素地塞米松诱导胸腺细胞凋亡作为模型系统，来研究凋亡过程中20 S和26 S蛋白酶体活性是否发生变化。我们观察到胸腺细胞含有高浓度的蛋白酶体，用地塞米松处理后，细胞提取物显示蛋白酶体类胰凝乳蛋白酶活性降低，这与观察到的凋亡程度相关。从凋亡胸腺细胞提取物中部分纯化出这些复合物后，20 S和26 S蛋白酶体的类胰凝乳蛋白酶活性降低仍然明显，因此这不是由于蛋白质周转率普遍增加导致的竞争所致。蛋白酶体复合物的类胰蛋白酶和肽基谷氨酰肽水解酶活性在凋亡过程中也被观察到降低，但这些降低通过半胱天冬酶抑制剂苄氧羰基-Val-Ala-Asp(OMe)-氟甲基酮抑制凋亡而被逆转。然而，在存在凋亡抑制剂的情况下，蛋白酶体的类胰凝乳蛋白酶活性进一步降低。发现Val-Ala-Asp-氟甲基酮在体外抑制26 S蛋白酶体的类胰凝乳蛋白酶和类胰蛋白酶活性。凋亡过程中蛋白酶体活性的降低似乎不是由于细胞内总蛋白酶体浓度的降低。因此，凋亡过程中20 S和26 S蛋白酶体活性的早期降低似乎是由于它们的蛋白水解活性下调，而不是由于它们的蛋白质浓度降低。这些数据表明，在胸腺细胞中，蛋白酶体可能负责一种作为凋亡正调节因子的蛋白质的周转。
