Disruption of the vacuolar-type H+-ATPase complex in liver causes MTORC1-independent accumulation of autophagic vacuoles and lysosomes
Disruption of the vacuolar-type H+-ATPase complex in liver causes MTORC1-independent accumulation of autophagic vacuoles and lysosomes
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DOI：
10.1080/15548627.2017.1280216
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发表时间：
2017-01-01
期刊：
影响因子：
13.3
通讯作者：
Saftig, Paul
中科院分区：
文献类型：
--
作者：
Kissing, Sandra;Rudnik, Soenke;Saftig, Paul
液泡型H+-转运ATP酶（v-H+-ATP酶）与雷帕霉素复合物1（MTORC 1）机制靶点的氨基酸依赖性激活有关，雷帕霉素复合物1是巨自噬的重要调节因子。为了揭示v-H+-ATP酶和MTORC 1之间的机制联系，我们通过诱导缺失必需的伴侣ATP 6AP 2来使小鼠肝细胞中的v-H+-ATP酶复合物去活性化。ATP 6AP 2突变体的特征在于在肝细胞中大量积累内吞和自噬空泡。这种细胞表型不是由内吞成熟阻滞或酸化受损引起的。然而，LC 3-II在敲除肝细胞中的降解似乎减少。当v-H+-ATP酶水平降低时，我们观察到尽管自噬标记蛋白增加，但MTOR与MTORC 1的正常信号传导的溶酶体关联。为了更好地理解为什么当v-H+-ATP酶耗尽时MTORC 1可以是活性的，在ATP 6AP 2缺陷型成纤维细胞中分析了MTORC 1的活化。在这些细胞中，观察到非常少的氨基酸引起的MTORC 1活化。相比之下，胰岛素诱导MTORC 1激活，这仍然需要细胞内的氨基酸储存。这些结果表明，在体内的宏观自噬的调节不仅依赖于V-H+-ATP酶介导的MTORC 1的调节。
The vacuolar-type H+-translocating ATPase (v-H+-ATPase) has been implicated in the amino aciddependent activation of the mechanistic target of rapamycin complex 1 (MTORC1), an important regulator of macroautophagy. To reveal the mechanistic links between the v-H+-ATPase and MTORC1, we destablilized v-H+-ATPase complexes in mouse liver cells by induced deletion of the essential chaperone ATP6AP2. ATP6AP2-mutants are characterized by massive accumulation of endocytic and autophagic vacuoles in hepatocytes. This cellular phenotype was not caused by a block in endocytic maturation or an impaired acidification. However, the degradation of LC3-II in the knockout hepatocytes appeared to be reduced. When v-H+-ATPase levels were decreased, we observed lysosome association of MTOR and normal signaling of MTORC1 despite an increase in autophagic marker proteins. To better understand why MTORC1 can be active when v-H+-ATPase is depleted, the activation of MTORC1 was analyzed in ATP6AP2-deficient fibroblasts. In these cells, very little amino acid-elicited activation of MTORC1 was observed. In contrast, insulin did induce MTORC1 activation, which still required intracellular amino acid stores. These results suggest that in vivo the regulation of macroautophagy depends not only on v-H+-ATPase-mediated regulation of MTORC1.