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Regulatory mechanism of autophagy and receptor trafficking by LAMP-2

LAMP-2对自噬和受体转运的调控机制

基本信息

批准号：
14580701
负责人：
TANAKA Yoshitaka
金额：
$ 1.98万
依托单位：
Kyushu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14580701/
关键词：
lysosome endosome lysosomal membrane protein autophagy protein sorting

项目摘要

We have generated mice deficient for lysosome-associated membrane protein-2 (LAMP-2) to investigate the physiological role of this protein. LAMP-2 deficiency leads to premature postnatal death of about. half of all LAMP-2 deficient mice. In several LAMP-2-deficient tissues, including muscle, heart, pancreas, and liver, an accumulation of autophagic vacuoles was observed. We also observed a reduced contractile function of the heart muscle. In this study, we extend the phenotype analysis using cultured hepatocytes. Enzyme activity measurements showed that the trafficking of some lysosomal enzymes to lysosomes was impaired. Immunoprecipitation of metabolically labeled cathepsin D indicated reduced intracellular retention and processing in the knockout cells. Interestingly, the steady-state level of 46-kDa mannose 6-phosphate receptor (MPR46) was decreased to 30% of controls due to a shorter half-life. Less receptor was found in the Golgi and in endosomes, suggesting impaired recycling from endosomes to the Golgi. More receptor was found in autophagic vacuoles, which may explain its shorter half-life. Our data indicate that in hepatocytes LAMP-2 deficiency either directly or indirectly leads to impaired recycling of MPR46 and partial mistargeting of a subset of lysosomal enzymes. Autophagic vacuoles may accumulale due to impaired capacity for lysosomal degradation.

我们已经产生了小鼠缺乏溶酶体相关膜蛋白-2（LAMP-2），以研究这种蛋白质的生理作用。LAMP-2缺乏导致大约100岁的早产儿死亡。所有LAMP-2缺陷小鼠的一半。在几种LAMP-2缺陷组织中，包括肌肉、心脏、胰腺和肝脏，观察到自噬空泡的积累。我们还观察到心肌收缩功能降低。在这项研究中，我们使用培养的肝细胞扩展表型分析。酶活性测定表明，一些溶酶体酶的运输到溶酶体受损。代谢标记的组织蛋白酶D的免疫沉淀表明敲除细胞中的细胞内滞留和加工减少。有趣的是，由于半衰期较短，46-kDa甘露糖6-磷酸受体（MPR 46）的稳态水平降低至对照组的30%。在高尔基体和内体中发现较少的受体，表明从内体到高尔基体的再循环受损。在自噬泡中发现较多的受体，这可能是其半衰期较短的原因。我们的数据表明，在肝细胞中，LAMP-2缺陷直接或间接导致MPR 46的再循环受损和溶酶体酶亚群的部分错误定位。由于溶酶体降解能力受损，自噬空泡可积聚。