Impaired turnover of autophagolysosomes in cathepsin L deficiency
Impaired turnover of autophagolysosomes in cathepsin L deficiency
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DOI：
10.1515/bc.2010.097
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发表时间：
2010-08-01
影响因子：
3.7
通讯作者：
Reinheckel, Thomas
中科院分区：
文献类型：
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作者：
Dennemaerker, Julia;Lohmueller, Tobias;Reinheckel, Thomas
溶酶体半胱氨酸内肽酶组织蛋白酶 L (Ctsl) 缺陷的小鼠的一些表型的特征是细胞质中存在大的变形囊泡。具体来说，心脏（扩张性心肌病）、甲状腺（甲状腺球蛋白加工受损）和角质形成细胞（周期性脱发和表皮过度增殖）受到影响。我们假设异常囊泡的形成是由于巨自噬的缺陷造成的。因此，我们研究了源自Ctsl(-/-)动物与自噬标记GFP-LC3转基因小鼠杂交的原代小鼠胚胎成纤维细胞(MEF)。 Ctsl(-/-) MEF 显示属于“酸性”细胞区室的囊泡结构的数量和大小增加，并且还具有 GFP-LC3 的特征。通过营养饥饿或雷帕霉素处理诱导自噬显示，Ctsl(-/-) MEF 中自噬的启动、自噬体的形成或自噬体-溶酶体融合没有明显受损，但 GFP-LC3 和 Lamp1 的共定位显示出异常大的充满 Lamp1 的自噬溶酶体。此外，可溶性溶酶体酶组织蛋白酶D在Ctsl(-/-)MEF中升高。因此，在没有 Ctsl 的情况下，自噬溶酶体内容物的降解会受到损害。这可能会减慢自噬溶酶体的周转，并导致之前在 Ctsl(-/-) 小鼠病理表型背景下描述的畸形和“酸性”囊泡的积累。
Some of the phenotypes of mice deficient for the lysosomal cysteine endopeptidase cathepsin L (Ctsl) are characterized by large dysmorphic vesicles in the cytoplasm. Specifically, the heart (dilative cardiomyopathy), the thyroid (impaired thyroglobulin processing) and keratinocytes (periodic hair loss and epidermal hyperproliferation) are affected. We hypothesized that the formation of aberrant vesicles is owing to defects in macroautophagy. Therefore, primary mouse embryonic fibroblasts (MEF), which were derived from Ctsl(-/-) animals crossed with mice transgenic for the autophagy marker GFP-LC3, were investigated. Ctsl(-/-) MEF show increased number and size of vesicular structures belonging to the 'acidic' cellular compartment and are also characterized by GFP-LC3. Induction of autophagy by nutrient starvation or rapamycin treatment showed no significant impairment of the initiation of autophagy, the formation of autophagosomes or autophagosome-lysosome fusion in Ctsl(-/-) MEF, but co-localization of GFP-LC3 and Lamp1 revealed unusually large autophagolysosomes filled with Lamp1. Furthermore, the soluble lysosomal enzyme cathepsin D was elevated in Ctsl(-/-) MEF. Thus, degradation of autophagolysosomal content is impaired in the absence of Ctsl. This could slow the turnover of autophagolysosomes and result in accumulation of the dysmorphic and 'acidic' vesicles that were previously described in the context of the pathological phenotypes of Ctsl(-/-) mice.