Morton P A, Zacheis M L, Giacoletto K S, Manning J A, Schwartz B D
Division of Immunology, G.D. Searle & Co., Chesterfield, MO 63198.
J Immunol. 1995 Jan 1;154(1):137-50.
The intracellular trafficking, proteolysis, and dissociation of invariant chain (li) associated with nascent class II molecules was examined in B-lymphoblastoid cells. Metabolic labeling and Percoll gradient centrifugation was used to assess the kinetics of delivery and processing of class II-li complexes within the endocytic pathway. Catabolism of class II-li complexes rapidly followed their delivery from post-Golgi compartments to dense lysosome-like compartments distinct from early and late endosomes. Direct peptide binding assays revealed that class II molecules associated with even small N-terminal fragments of li failed to bind peptide. Cysteine protease inhibitors alone blocked li proteolysis/dissociation and accumulation of class II-li biosynthetic intermediates within lysosome-containing compartments. Active-site labeling of cysteine proteases in B cells was used to identify cysteine proteases capable of mediating li proteolysis within endosomal compartments. Our results indicate rapid, possibly direct, transport of nascent class II-li complexes from the Golgi/trans-Golgi network to dense lysosomal compartments wherein cysteine protease(s), likely including cathepsin B, mediate complete removal of li. Inhibition of cysteine protease activity results in the accumulation of incompletely processed class II-li complexes, which lack peptide binding ability, within lysosomal compartments.
在B淋巴母细胞中研究了与新生II类分子相关的恒定链（Ii）的细胞内运输、蛋白水解和解离。采用代谢标记和Percoll梯度离心法评估内吞途径中II类-Ii复合物的递送和加工动力学。II类-Ii复合物从高尔基体后区室递送至与早期和晚期内体不同的致密溶酶体样区室后，迅速发生分解代谢。直接肽结合试验表明，与Ii的小N端片段相关的II类分子无法结合肽。单独使用半胱氨酸蛋白酶抑制剂可阻断Ii的蛋白水解/解离以及含溶酶体区室内II类-Ii生物合成中间体的积累。通过对B细胞中半胱氨酸蛋白酶进行活性位点标记，以鉴定能够在内体区室中介导Ii蛋白水解的半胱氨酸蛋白酶。我们的结果表明，新生的II类-Ii复合物从高尔基体/反式高尔基体网络快速运输至致密溶酶体区室，其中半胱氨酸蛋白酶（可能包括组织蛋白酶B）介导Ii的完全去除。抑制半胱氨酸蛋白酶活性会导致含溶酶体区室内缺乏肽结合能力的未完全加工的II类-Ii复合物积累。
