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Published on: April 24, 2021
Inhibition of ESCRT-III activates alternative pathways for protein degradation and secretion
Masaru Fujiwara1, Ryohei Sakai2, Masayuki Takahashi2
1Department of Degenerative Neurological Diseases, National Institute of Neuroscience, National Center of Neurology and Psychiatry, 4-1-1 Ogawa-Higashi, Kodaira, Tokyo, 187-8502, Japan; Department of Biotechnology and Life Science, Faculty of Technology, Tokyo University of Agriculture and Technology, 2-24-16 Naka-machi, Koganei, Tokyo, 184-8588, Japan.
Abstract:
Mutations in components of the endosomal sorting complex required for transport (ESCRT)-III, such as CHMP2B and VPS4A/B, are known to cause neurological disorders, including frontotemporal lobar degeneration (FTLD) and developmental encephalopathies. Although ESCRT complexes are required for macroautophagy and for certain forms of microautophagy, the effects of ESCRT-III dysfunction on intracellular protein degradation remain unclear. In this study, we investigated how ESCRT-III dysfunction affects intracellular protein clearance using multiple genetic manipulations, including a dominant-negative form of VPS4 and an FTLD-associated CHMP2B mutant. We found that despite marked suppression of macroautophagic flux, inhibition of ESCRT-III promoted protein clearance in multiple cell types, including Neuro2a cells. Such clearance was also observed in ATG13-or ATG5-knockout cells, confirming that this process occurs independently of macroautophagy. Imaging revealed increased punctate accumulation of substrate proteins in lysosomes, suggesting the activation of a microautophagy-like pathway independent of ESCRT-III. In addition, ESCRT-III inhibition enhances extracellular vesicle-independent protein secretion. Cell-to-cell transmission of aggregated tau, assessed using conditioned medium, was also promoted by ESCRT-III inhibition. These findings suggested that ESCRT-III dysfunction, while impairing canonical autophagy, paradoxically activates alternative degradation and secretion pathways that may contribute to the pathogenesis of neurological disorders.
Insights
Endosomal sorting complex (ESCRT)-III dysfunction impairs autophagy but enhances protein clearance via alternative pathways. This unexpected clearance, independent of macroautophagy, may contribute to neurological disease pathogenesis.
Area of Science:
- Cell Biology
- Neuroscience
- Molecular Biology
Background:
- Mutations in ESCRT-III components (e.g., CHMP2B, VPS4A/B) are linked to neurological disorders like FTLD.
- ESCRT-III's role in intracellular protein degradation, particularly its impact on autophagy, remains incompletely understood.
Purpose of the Study:
- To investigate the effects of ESCRT-III dysfunction on intracellular protein clearance mechanisms.
- To determine if ESCRT-III inhibition impacts autophagy and other protein degradation pathways.
Main Methods:
- Genetic manipulation using a dominant-negative VPS4 form and an FTLD-associated CHMP2B mutant.
- Assessment of protein clearance in various cell types, including ATG13/ATG5-knockout cells.
- Analysis of lysosomal substrate accumulation and extracellular vesicle-independent protein secretion.
Main Results:
- ESCRT-III inhibition suppressed macroautophagic flux but paradoxically promoted protein clearance.
- Protein clearance occurred independently of macroautophagy, suggesting alternative pathways.
- Increased lysosomal accumulation of substrate proteins indicated microautophagy activation.
- Enhanced extracellular vesicle-independent protein secretion and cell-to-cell tau transmission were observed.
Conclusions:
- ESCRT-III dysfunction impairs canonical autophagy but activates alternative protein degradation and secretion pathways.
- These alternative pathways, including microautophagy-like processes, may play a role in the pathogenesis of neurological disorders.
- Understanding these compensatory mechanisms is crucial for developing therapeutic strategies for ESCRT-III-related diseases.
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