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RAB7A GTPase Is Involved in Mitophagosome Formation and Autophagosome–Lysosome Fusion in N2a Cells Treated with the Prion Protein Fragment 106–126

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Abstract

Failed communication between mitochondria and lysosomes causes dysfunctional mitochondria, which may induce mitochondria-related neurodegenerative diseases. Here, we show that RAB7A, a small GTPase of the Rab family, mediates the crosstalk between these two important organelles to maintain homeostasis in N2a cells treated with PrP106–126. Specifically, we demonstrate that mitophagy deficiency in N2a cells caused by PrP106–126 is associated with dysregulated RAB7A localization in mitochondria. Cells lacking RAB7A display decreased mitochondrial colocalization with lysosomes and significantly increased mitochondrial protein expression, resulting in inhibited mitophagy. In contrast, overexpression of GTP-bound RAB7A directly induces lysosome colocalization with mitochondria. Further study revealed that GTP-bound RAB7A protects mitochondrial homeostasis by supporting autophagosome biogenesis. Moreover, we suggest that depletion of RAB7A leads to gross morphological changes in lysosomes, which prevents autophagosome–lysosome fusion and interferes with the breakdown of autophagic cargo within lysosomes. Overexpression of GTP-bound RAB7A can also alleviate PrP106–126-induced morphological damage and dysfunction of mitochondria, reducing neuronal apoptosis. Collectively, our data demonstrate that RAB7A successfully drives mitochondria to the autophagosomal lumen for degradation, suggesting that the communication of proteotoxic stress from mitochondria to lysosomes requires RAB7A, as a signaling molecule, to establish a link between the disturbed mitochondrial network and its remodeling. These findings indicate that small molecules regulating mitophagy have the potential to modulate cellular homeostasis and the clinical course of neurodegenerative diseases.

Graphical Abstract

Proposed model of mitophagy regulated by RAB7A. (1) Accumulating PrP106–126 induced mitophagy. (2) RAB7A is recruited to mitochondria. (3) ATG5-12 and ATG9A (5) vesicles are recruited to the autophagosome formation sites in a RAB7A-dependent manner. The ATG5-12 complex recruits and anchors LC3-I to form active LC3-II (4), accelerating mitophagosomal formation. The ATG9A vesicles are thought to be a source of membranes for autophagosome assembly. The recruitment of proteins and lipids induces membrane expansion and subsequent closure to form the mitophagosome. (6) Maintenance of the normal low lysosomal PH depends on active (GTP-bound) RAB7A. (7) RAB7A recruits effector molecules responsible for tight membrane interactions, and directly or indirectly, the subsequent autophagosome merges with the lysosome, and the cargo is completely degraded.

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Data Availability

Most of the data generated or analyzed during this study are included in the published article and its supplementary information files. Additional generated datasets are available from the corresponding author on reasonable request.

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Acknowledgements

We thank all the Prion Laboratory members for technical support and input. We are grateful to Prof. Qiao Jian and Lifeng Yang for helpful ideas and experimental design. We also thank Prof. Jianxin Chen of South China Agricultural University for manuscript revisions.

Funding

This work was supported by the Natural Science Foundation of China (Project No.31972641).

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Authors and Affiliations

  1. National Animal Transmissible Spongiform Encephalopathy Laboratory, College of Veterinary Medicine, China Agricultural University, Beijing, 100193, China

    Zhiping Li, Mengyu Lai, Jie Li, Dongming Yang, Mengyang Zhao, Dongdong Wang, Zhixin Sun, Pei Wen, Fengting Gou, Yuexin Dai, Yilan Ji, Deming Zhao & Lifeng Yang

  2. Department of Pathophysiology, College of Veterinary Medicine, China Agricultural University, Beijing, 100193, China

    Zhiping Li & Jian Qiao

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  1. Zhiping Li
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Contributions

ZL and LY conceived and designed the experiments. ZL performed the experiments, and ZL interpreted the results. ZL, JQ, and LY contributed to manuscript writing. LY reviewed and revised the paper. All authors read and approved the final manuscript.

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Correspondence to Jian Qiao or Lifeng Yang.

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Li, Z., Lai, M., Li, J. et al. RAB7A GTPase Is Involved in Mitophagosome Formation and Autophagosome–Lysosome Fusion in N2a Cells Treated with the Prion Protein Fragment 106–126. Mol Neurobiol 60, 1391–1407 (2023). https://doi.org/10.1007/s12035-022-03118-5

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