Changes in the proteolytic activities of proteasomes and lysosomes in human fibroblasts produced by serum withdrawal, amino-acid deprivation and confluent conditions
Changes in the proteolytic activities of proteasomes and lysosomes in human fibroblasts produced by serum withdrawal, amino-acid deprivation and confluent conditions
复制标题
DOI：
10.1042/bj20030282
复制
发表时间：
2003-10-01
影响因子：
4.1
通讯作者：
Knecht, E
中科院分区：
文献类型：
--
作者：
Fuertes, G;De Llano, JJM;Knecht, E
研究了在不同条件下生长的人成纤维细胞中主要的蛋白质水解途径对长寿命蛋白质降解的贡献。首先详细分析了各种常用的蛋白质降解药理抑制剂的作用。通过选择溶酶体和蛋白酶体的特定抑制剂。据观察，这两种途径共同占细胞蛋白质降解的80%或更多。对于溶酶体抑制剂，发现血清停药或氨基酸剥夺强烈刺激巨噬，但对其他溶酶体途径没有作用，而融合条件对巨噬没有影响，并轻微激活其他溶酶体途径。长时间（24 h）血清饥饿显著降低了巨噬途径，而其他溶酶体途径的活性增加。这些变化与溶酶体的电镜观察和形态测量相关。使用蛋白酶体抑制剂。人们发现。在没有血清的情况下呈指数增长的细胞中，泛素-蛋白酶体途径的活性增加，而在融合条件下，蛋白酶体对降解的贡献（以百分比计）减少，特别是在缺乏氨基酸的细胞中。有趣的是，在融合细胞中，19 S调节复合体的两种成分和可互换的β亚基的水平下降。这与pa28免疫蛋白酶体成分水平的显著增加有关。因此，汇合性疾病影响蛋白酶体的方式类似于干扰素治疗。总之，这些结果表明，各种蛋白水解途径的活性取决于细胞的生长条件，并将有助于研究控制其活性的特定信号。
The contribution of the main proteolytic pathways to the degradation of long-lived proteins in human fibroblasts grown under different conditions was investigated. The effects of various commonly used pharmacological inhibitors of protein degradation were first analysed in detail. By choosing specific inhibitors of lysosomes and proteasomes. it was observed that together both pathways accounted for 80 % or more of the degradation of cell proteins. With lysosomal inhibitors, it was found that serum withdrawal or amino-acid deprivation strongly stimulated macroautophagy but not other lysosomal pathways, whereas confluent conditions had no effect on macroautophagy and slightly activated other lysosomal pathways. Prolonged (24 h) serum starvation of confluent cultures strongly decreased the macroautophagic pathway, whereas the activity of other lysosomal pathways increased. These changes correlated with electron microscopic observations and morphometric measurements of lysosomes. With proteasomal inhibitors. it was found that. in exponentially growing cells in the absence of serum, activity of the ubiquitin-proteasome pathway increases, whereas under confluent conditions the contribution (in percentage) of proteasomes to degradation decreases, especially in cells deprived of amino acids. Interestingly, in confluent cells, the levels of two components of the 19 S regulatory complex and those of an interchangeable beta-subunit decreased. This was associated with a marked increase in the levels of components of PA28-immunoproteasomes. Thus confluent conditions affect proteasomes in a way that resembles treatment with interferon-gamma. Altogether, these results show that the activity of the various proteolytic pathways depends on the growth conditions of cells and will be useful for investigation of the specific signals that control their activity.