Hopgood M F, Knowles S E, Ballard F J
Biomed Biochim Acta. 1986;45(11-12):1603-10.
Several glycolytic enzymes as well as horseradish peroxidase have been labelled by reductive methylation and microinjected into rat L6 myoblasts using the erythrocyte ghost fusion technique. Since endogenous protein in the recipient cells was labelled with [14C]leucine, the degradation of both injected and endogenous protein could be followed simultaneously. The experiments show that inhibitors of autophagy generally affect the catabolism of total endogenous protein to a greater extent than the injected proteins. Both lactate dehydrogenase and peroxidase are poorly susceptible to the inhibitors even though these two enzymes represent virtually the most and least stable, respectively, of the test proteins. Accordingly the results do not support the hypothesis that autophagy contributes more to the turnover of long half-life proteins. However, this interpretation should be treated cautiously because some of the injected proteins do not appear to be targeted to the cell cytosol by the fusion process, and the breakdown of such proteins is only poorly susceptible to inhibition by insulin, serum or cycloheximide.
几种糖酵解酶以及辣根过氧化物酶已通过还原甲基化进行标记，并使用红细胞血影融合技术显微注射到大鼠L6成肌细胞中。由于受体细胞中的内源性蛋白质用[14C]亮氨酸进行了标记，因此可以同时追踪注射蛋白和内源性蛋白的降解情况。实验表明，自噬抑制剂通常对内源性总蛋白分解代谢的影响比对注射蛋白的影响更大。乳酸脱氢酶和过氧化物酶对抑制剂的敏感性都很低，尽管这两种酶实际上分别代表了测试蛋白中最稳定和最不稳定的。因此，结果不支持自噬对长半衰期蛋白周转贡献更大的假设。然而，这种解释应谨慎对待，因为一些注射蛋白似乎没有通过融合过程靶向到细胞胞质溶胶，并且此类蛋白的分解对胰岛素、血清或环己酰亚胺抑制的敏感性很低。
