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.

我们已经产生了缺乏与溶酶体相关的膜蛋白-2（LAMP-2）的小鼠研究该蛋白的生理作用。 LAMP-2缺乏会导致大约早产后死亡。所有Lamp-2不足小鼠的一半。在包括肌肉，心脏，胰腺和肝脏在内的几种LAMP-2-2缺陷组织中，观察到自噬液泡的积累。我们还观察到心肌的收缩功能降低。在这项研究中，我们使用培养的肝细胞扩展了表型分析。酶活性测量表明，某些溶酶体酶向溶酶体的运输受损。代谢标记的组织蛋白酶D的免疫沉淀表明敲除细胞中细胞内保留和加工降低。有趣的是，由于半衰期较短，46 kDa甘露糖6-磷酸受体（MPR46）的稳态水平降低到30％。在高尔基体和内体中发现的受体较少，表明从内体到高尔基体的回收受损。在自噬液泡中发现了更多的受体，这可以解释其较短的半衰期。我们的数据表明，在肝细胞LAMP-2缺乏症中，直接或间接导致MPR46的回收障碍和溶酶体酶子集的部分误射。自噬液泡可能由于溶酶体降解的能力受损而累积。