The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup>
Neuronal oxidative stress caused by mitochondrial dysfunction plays a crucial role in the development of Parkinson’s disease (PD). Growing evidence shows that autophagy confers neuroprotection in oxidative-stress-associated PD. This work aims to investigate the involvement of TMEM166, an endoplasmic...
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2022-08-01
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| author | Zhaozhong Liao Zunshuang Gong Zhe Wang Weiyan Yang Wenjing Liu Lin Hou Xiaokun Liu Junnan Hua Bin Wang Ning Li |
| author_facet | Zhaozhong Liao Zunshuang Gong Zhe Wang Weiyan Yang Wenjing Liu Lin Hou Xiaokun Liu Junnan Hua Bin Wang Ning Li |
| author_sort | Zhaozhong Liao |
| collection | DOAJ |
| description | Neuronal oxidative stress caused by mitochondrial dysfunction plays a crucial role in the development of Parkinson’s disease (PD). Growing evidence shows that autophagy confers neuroprotection in oxidative-stress-associated PD. This work aims to investigate the involvement of TMEM166, an endoplasmic-reticulum-localized autophagy-regulating protein, in the process of PD-associated oxidative stress through the classic cellular PD model of neuroblastoma SH-SY5Y cells exposed to 1-methyl-4-phenylpyridinium (MPP<sup>+</sup>). Reactive oxygen species (ROS) production and mitochondrial membrane potential were checked to assess the oxidative stress induced by MPP<sup>+</sup> and the cellular ATP generated was determined to evaluate mitochondrial function. The effect on autophagy induction was evaluated by analyzing p62 and LC3-II/I expression and by observing the LC3 puncta and the colocalization of LC3 with LAMP1/ LAMP2. The colocalization of mitochondria with LC3, the colocalization of Tom20 with LAMP1 and Tom20 expression were analyzed to evaluate mitophagy. We found that TMEM166 is up-regulated in transcript levels, but up-regulated first and then down-regulated by autophagic degradation in protein levels upon MPP<sup>+</sup>-treatment. Overexpression of TMEM166 induces mitochondria fragmentation and dysfunction and exacerbates MPP<sup>+</sup>-induced oxidative stress and cell viability reduction. Overexpression of TMEM166 is sufficient to induce autophagy and mitophagy and promotes autophagy and mitophagy under MPP<sup>+</sup> treatment, while knockdown of TMEM166 inhibits basal autophagic degradation. In addition, overexpressed TMEM166 suppresses AMPK activation, while TMEM166 knockdown enhances AMPK activation. Pharmacological activation of AMPK alleviates the exacerbation of oxidative stress induced by TMEM166 overexpression and increases cell viability, while pharmacological inhibition mitophagy aggravates the oxidative stress induced by MPP<sup>+</sup> treatment combined with TMEM166 overexpression. Finally, we find that overexpressed TMEM166 partially localizes to mitochondria and, simultaneously, the active AMPK in mitochondria is decreased. Collectively, these findings suggest that TMEM166 can translocate from ER to mitochondria and inhibit AMPK activation and, in response to mitochondrial oxidative stress, neuronal cells choose to up-regulate TMEM166 to promote autophagy/mitophagy; then, the enhancing autophagy/mitophagy degrades the TMEM166 to activate AMPK, by the two means to maintain cell survival. The continuous synthesis and degradation of TMEM166 in autophagy/mitochondria flux suggest that TMEM166 may act as an autophagy/mitochondria adaptor. |
| first_indexed | 2024-03-10T01:57:25Z |
| format | Article |
| id | doaj.art-f5fa9f592cf0411097cfe26238e4c44a |
| institution | Directory Open Access Journal |
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| language | English |
| last_indexed | 2024-03-10T01:57:25Z |
| publishDate | 2022-08-01 |
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| spelling | doaj.art-f5fa9f592cf0411097cfe26238e4c44a2023-11-23T12:55:36ZengMDPI AGCells2073-44092022-08-011117270610.3390/cells11172706The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup>Zhaozhong Liao0Zunshuang Gong1Zhe Wang2Weiyan Yang3Wenjing Liu4Lin Hou5Xiaokun Liu6Junnan Hua7Bin Wang8Ning Li9Department of Biochemistry and Molecular Biology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaDepartment of Biochemistry and Molecular Biology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaDepartment of Biotechnology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaDepartment of Anesthesiology, Family Planning Service Center, Maternal and Child Health Hospital of Jiaozhou City, Qingdao 266000, ChinaDepartment of Biochemistry and Molecular Biology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaDepartment of Biochemistry and Molecular Biology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaDepartment of Biochemistry and Molecular Biology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaDepartment of Biotechnology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaCollege of Electronic Information, Micro-Nano Technology College, Qingdao University, Qingdao 266000, ChinaDepartment of Biochemistry and Molecular Biology, School of Basic Medicine, Qingdao University, Qingdao 266000, ChinaNeuronal oxidative stress caused by mitochondrial dysfunction plays a crucial role in the development of Parkinson’s disease (PD). Growing evidence shows that autophagy confers neuroprotection in oxidative-stress-associated PD. This work aims to investigate the involvement of TMEM166, an endoplasmic-reticulum-localized autophagy-regulating protein, in the process of PD-associated oxidative stress through the classic cellular PD model of neuroblastoma SH-SY5Y cells exposed to 1-methyl-4-phenylpyridinium (MPP<sup>+</sup>). Reactive oxygen species (ROS) production and mitochondrial membrane potential were checked to assess the oxidative stress induced by MPP<sup>+</sup> and the cellular ATP generated was determined to evaluate mitochondrial function. The effect on autophagy induction was evaluated by analyzing p62 and LC3-II/I expression and by observing the LC3 puncta and the colocalization of LC3 with LAMP1/ LAMP2. The colocalization of mitochondria with LC3, the colocalization of Tom20 with LAMP1 and Tom20 expression were analyzed to evaluate mitophagy. We found that TMEM166 is up-regulated in transcript levels, but up-regulated first and then down-regulated by autophagic degradation in protein levels upon MPP<sup>+</sup>-treatment. Overexpression of TMEM166 induces mitochondria fragmentation and dysfunction and exacerbates MPP<sup>+</sup>-induced oxidative stress and cell viability reduction. Overexpression of TMEM166 is sufficient to induce autophagy and mitophagy and promotes autophagy and mitophagy under MPP<sup>+</sup> treatment, while knockdown of TMEM166 inhibits basal autophagic degradation. In addition, overexpressed TMEM166 suppresses AMPK activation, while TMEM166 knockdown enhances AMPK activation. Pharmacological activation of AMPK alleviates the exacerbation of oxidative stress induced by TMEM166 overexpression and increases cell viability, while pharmacological inhibition mitophagy aggravates the oxidative stress induced by MPP<sup>+</sup> treatment combined with TMEM166 overexpression. Finally, we find that overexpressed TMEM166 partially localizes to mitochondria and, simultaneously, the active AMPK in mitochondria is decreased. Collectively, these findings suggest that TMEM166 can translocate from ER to mitochondria and inhibit AMPK activation and, in response to mitochondrial oxidative stress, neuronal cells choose to up-regulate TMEM166 to promote autophagy/mitophagy; then, the enhancing autophagy/mitophagy degrades the TMEM166 to activate AMPK, by the two means to maintain cell survival. The continuous synthesis and degradation of TMEM166 in autophagy/mitochondria flux suggest that TMEM166 may act as an autophagy/mitochondria adaptor.https://www.mdpi.com/2073-4409/11/17/2706TMEM166EVA1Aoxidative stressParkinson’s diseasemitochondriaAMPK |
| spellingShingle | Zhaozhong Liao Zunshuang Gong Zhe Wang Weiyan Yang Wenjing Liu Lin Hou Xiaokun Liu Junnan Hua Bin Wang Ning Li The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup> Cells TMEM166 EVA1A oxidative stress Parkinson’s disease mitochondria AMPK |
| title | The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup> |
| title_full | The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup> |
| title_fullStr | The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup> |
| title_full_unstemmed | The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup> |
| title_short | The Degradation of TMEM166 by Autophagy Promotes AMPK Activation to Protect SH-SY5Y Cells Exposed to MPP<sup>+</sup> |
| title_sort | degradation of tmem166 by autophagy promotes ampk activation to protect sh sy5y cells exposed to mpp sup sup |
| topic | TMEM166 EVA1A oxidative stress Parkinson’s disease mitochondria AMPK |
| url | https://www.mdpi.com/2073-4409/11/17/2706 |
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