Carbohydrates act as sorting determinants in ER-associated degradation of tyrosinase
Carbohydrates act as sorting determinants in ER-associated degradation of tyrosinase
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DOI：
10.1242/jcs.01154
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发表时间：
2004-06-15
影响因子：
4
通讯作者：
Hebert, DN
中科院分区：
文献类型：
--
作者：
Svedine, S;Wang, T;Hebert, DN
内质网（ER）质量控制机制通过对异常蛋白质进行内质网相关蛋白质降解（ERAD）来维持成熟过程的准确性，这一过程需要从内质网腔逆向转运到细胞质，并由蛋白酶体进行降解。在此，我们通过监测野生型（Tyr）和白化突变型（Tyr(C85S)）酪氨酸酶的降解来评估N - 连接聚糖在内质网相关蛋白质降解中的作用。首先，利用完整的黑素细胞和半透性细胞系统，确定突变型酪氨酸酶是一种真正的内质网相关蛋白质降解底物。抑制甘露糖修剪或将Tyr(C85S)以单葡萄糖基化形式积累会使其稳定，这支持了凝集素伴侣在内质网滞留和蛋白酶体降解中的作用。相反，通过阻止葡萄糖修剪来消除凝集素伴侣相互作用会导致酪氨酸酶迅速消失，最初是由于蛋白质聚集体的形成，随后这些聚集体被蛋白酶体降解。聚集的酪氨酸酶与蛋白质二硫键异构酶和BiP共定位，但不与钙连蛋白共定位，这支持了一种内质网组织形式，它有助于蛋白质的成熟和降解。基于这些研究，我们提出了一个酪氨酸酶降解模型，其中N - 连接聚糖与凝集素伴侣之间的相互作用有助于减少酪氨酸酶聚集，并将非天然底物靶向逆向转运及随后的降解。
The endoplasmic reticulum (ER) quality-control machinery maintains the fidelity of the maturation process by sorting aberrant proteins for ER-associated protein degradation (ERAD), a process requiring retrotranslocation from the ER lumen to the cytosol and degradation by the proteasome. Here, we assessed the role of N-linked glycans in ERAD by monitoring the degradation of wild-type (Tyr) and albino mutant (Tyr(C85S)) tyrosinase. Initially, mutant tyrosinase was established as a genuine ERAD substrate using intact melanocyte and semi-permeabilized cell systems. Inhibiting mannose trimming or accumulating Tyr(C85S) in a monoglucosylated form led to its stabilization, supporting a role for lectin chaperones in ER retention and proteasomal degradation. In contrast, ablating the lectin chaperone interactions by preventing glucose trimming caused a rapid disappearance of tyrosinase, initially due to the formation of protein aggregates, which were subsequently degraded by the proteasome. The colocalization of aggregated tyrosinase with protein disulfide isomerase and BiP, but not calnexin, supports an ER organization, which aids in protein maturation and degradation. Based on these studies, we propose a model of tyrosinase degradation in which interactions between N-linked glycans and lectin chaperones help to minimize tyrosinase aggregation and also target non-native substrates for retro-translocation and subsequent degradation.