Mainferme F, Wattiaux R, von Figura K
Eur J Biochem. 1985 Nov 15;153(1):211-6. doi: 10.1111/j.1432-1033.1985.tb09288.x.
The synthesis, transport and processing of cathepsin C was studied in Morris hepatoma 7777 cells by metabolic labelling, immunoprecipitation and characterization of labelled polypeptides by gel electrophoresis and fluorography. The largest detectable precursor of cathepsin C was a polypeptide of Mr = 92 500. Even 3 min after synthesis this precursor was accompanied by four polypeptides with Mr values ranging from 63 000 to 54 000, indicating cleavage of the precursors within the endoplasmic reticulum. The early forms of cathepsin C were associated with low-buoyant-density organelles containing the markers of endoplasmic reticulum and Golgi complex. About 30% of these early forms were secreted within 3 h after synthesis. The remaining 70% were transferred into dense lysosomes and processed between 2 and 3 h after synthesis to a mixture of the least five major and nine minor polypeptides with Mr values ranging from 73 000 to 12 000. These forms remained stable for at least 3 days. In freshly isolated hepatocytes cathepsin C was processed to forms closely related to those found in the hepatoma cells. Cathepsin C was synthesized in Morris hepatoma 7777 cells as a glycoprotein with mannose-6-phosphate residues that mediated mannose-6-phosphate-specific receptor-dependent uptake in human skin fibroblasts. In contrast to hepatocytes, synthesis of mannose-6-phosphate receptors in Morris hepatoma 7777 cells was below the limit of detection. The hepatoma cells did not express at the cell surface these or other receptors mediating endocytosis of lysosomal enzymes. Further, processing and transport of newly synthesized cathepsin C was largely resistant to NH4Cl. Apparently, cathepsin C is transferred in Morris hepatoma 7777 cells by a mechanism independent of mannose-6-phosphate-specific receptors.
通过代谢标记、免疫沉淀以及凝胶电泳和荧光自显影对标记多肽进行表征，研究了组织蛋白酶C在莫里斯肝癌7777细胞中的合成、转运和加工过程。组织蛋白酶C可检测到的最大前体是一种分子量为92500的多肽。即使在合成后3分钟，该前体也伴随着四种分子量在63000至54000之间的多肽，这表明前体在内质网内发生了切割。组织蛋白酶C的早期形式与含有内质网和高尔基体标志物的低浮力密度细胞器相关。这些早期形式中约30%在合成后3小时内被分泌。其余70%被转运至致密溶酶体，并在合成后2至3小时内加工成至少五种主要和九种次要多肽的混合物，其分子量在73000至12000之间。这些形式至少稳定3天。在新鲜分离的肝细胞中，组织蛋白酶C被加工成与肝癌细胞中发现的形式密切相关的形式。组织蛋白酶C在莫里斯肝癌7777细胞中作为一种含有甘露糖-6-磷酸残基的糖蛋白合成，这些残基介导了在人皮肤成纤维细胞中依赖甘露糖-6-磷酸特异性受体的摄取。与肝细胞相反，莫里斯肝癌7777细胞中甘露糖-6-磷酸受体的合成低于检测限。肝癌细胞在细胞表面不表达这些或其他介导溶酶体酶内吞作用的受体。此外，新合成的组织蛋白酶C的加工和转运在很大程度上对氯化铵具有抗性。显然，组织蛋白酶C在莫里斯肝癌7777细胞中的转运机制独立于甘露糖-6-磷酸特异性受体。
