Chaperone-mediated autophagy protects against hyperglycemic stress

by Selleckchem | Dec 12 2023

Chaperone-mediated autophagy (CMA) is a major pathway of lysosomal proteolysis critical for cellular homeostasis and metabolism, and whose defects have been associated with several human pathologies. While CMA has been well described in mammals, functional evidence has only recently been documented in fish, opening up new perspectives to tackle this function under a novel angle. Now we propose to explore CMA functions in the rainbow trout (RT, Oncorhynchus mykiss), a fish species recognized as a model organism of glucose intolerance and characterized by the presence of two paralogs of the CMA-limiting factor Lamp2A (lysosomal associated membrane protein 2A). To this end, we validated a fluorescent reporter (KFERQ-PA-mCherry1) previously used to track functional CMA in mammalian cells, in an RT hepatoma-derived cell line (RTH-149). We found that incubation of cells with high-glucose levels (HG, 25 mM) induced translocation of the CMA reporter to lysosomes and/or late endosomes in a KFERQ- and Lamp2A-dependent manner, as well as reduced its half-life compared to the control (5 mM), thus demonstrating increased CMA flux. Furthermore, we observed that activation of CMA upon HG exposure was mediated by generation of mitochondrial reactive oxygen species, and involving the antioxidant transcription factor Nfe2l2/Nrf2 (nfe2 like bZIP transcription factor 2). Finally, we demonstrated that CMA plays an important protective role against HG-induced stress, primarily mediated by one of the two RT Lamp2As. Together, our results provide unequivocal evidence for CMA activity existence in RT and highlight both the role and regulation of CMA during glucose-related metabolic disorders.

Comments:

The passage you provided describes a scientific study exploring chaperone-mediated autophagy (CMA) in rainbow trout (RT), a fish species known for its relevance as a model organism for glucose intolerance. Here's a breakdown of the key findings and concepts mentioned in the passage:

1. **Background**: Chaperone-mediated autophagy (CMA) is a vital cellular process responsible for lysosomal proteolysis, crucial for maintaining cellular homeostasis and metabolism. Dysfunctions in CMA have been associated with various human diseases.

2. **Study Focus**: The study aimed to investigate CMA functions in rainbow trout, a fish species with known glucose intolerance, by utilizing a fluorescent reporter (KFERQ-PA-mCherry1) previously used in mammalian cells to track functional CMA.

3. **Experimental Setup**: An RT hepatoma-derived cell line (RTH-149) was used for the experiments. The cells were exposed to high-glucose levels (25 mM) to simulate conditions of glucose intolerance.

4. **Key Findings**:
    - **CMA Activation**: High-glucose exposure induced translocation of the CMA reporter to lysosomes and/or late endosomes in a manner dependent on the specific CMA-targeting motif KFERQ and the CMA-limiting factor Lamp2A. This indicated an increased CMA flux under high-glucose conditions.
    - **Regulation by Reactive Oxygen Species (ROS)**: CMA activation in response to high-glucose levels was mediated by the generation of mitochondrial reactive oxygen species (ROS). ROS are chemically reactive molecules containing oxygen, often produced as byproducts of cellular metabolism.
    - **Involvement of Nfe2l2/Nrf2**: The activation of CMA in response to high-glucose conditions involved the antioxidant transcription factor Nfe2l2/Nrf2. Nrf2 plays a role in cellular response to oxidative stress.
    - **Protective Role**: CMA was found to have a significant protective role against high-glucose-induced stress, primarily mediated by one of the two Lamp2A paralogs present in rainbow trout.

5. **Significance**: The study provides clear evidence for the existence of CMA activity in rainbow trout and sheds light on the role and regulation of CMA in the context of glucose-related metabolic disorders. This research contributes to our understanding of cellular mechanisms in fish species and may have implications for understanding similar processes in other organisms, potentially including humans.

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