Autophagosome membrane expansion is mediated by the N-terminus and cis-membrane association of human ATG8s
- The Francis Crick Institute, United Kingdom
- University of Tokyo, Japan
- CSIR-Institute of Genomics and Integrative Biology, India
Abstract
Autophagy is an essential catabolic pathway which sequesters and engulfs cytosolic substrates via autophagosomes, unique double-membraned structures. ATG8 proteins are ubiquitin-like proteins recruited to autophagosome membranes by lipidation at the C-terminus. ATG8s recruit substrates, such as p62, and play an important role in mediating autophagosome membrane expansion. However, the precise function of lipidated ATG8 in expansion remains obscure. Using a real-time in vitro lipidation assay, we revealed that the N-termini of lipidated human ATG8s (LC3B and GABARAP) are highly dynamic and interact with the membrane. Moreover, atomistic MD simulation and FRET assays indicate that N-termini of LC3B and GABARAP associate in cis on the membrane. By using non-tagged GABARAPs, we show that GABARAP N-terminus and its cis-membrane insertion are crucial to regulate the size of autophagosomes in cells irrespectively of p62 degradation. Our study provides fundamental molecular insights into autophagosome membrane expansion, revealing the critical and unique function of lipidated ATG8.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting file; Source Data files have been provided for Figures 1 - 6.
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Funding
European Research Council (FP7/2007-2013 788708)
- Wenxin Zhang
- Sharon A Tooze
Japan Science and Technology, ERATO (JPMJER1702)
- Chieko Saito
Council of Scientific and Industrial Research, India (OLP1163)
- Deepanshi Gahlot
- Lipi Thukral
Wellcome Trust (CC2134)
- Wenxin Zhang
- Harold BJ Jefferies
- Sharon A Tooze
Wellcome Trust (CC2064)
- Colin Davis
- Anne Schreiber
Cancer Research UK (CC2134)
- Wenxin Zhang
- Harold BJ Jefferies
- Sharon A Tooze
Cancer Research UK (CC2064)
- Colin Davis
- Anne Schreiber
Medical Research Council (CC2134)
- Wenxin Zhang
- Harold BJ Jefferies
- Sharon A Tooze
Medical Research Council (CC2064)
- Colin Davis
- Anne Schreiber
Japan Science and Technology, PRESTO (JPMJPR20EC)
- Taki Nishimura
Grant-in-Aid for Transformative Research Areas (B (grant 21H05146)
- Taki Nishimura
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2023, Zhang et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Autophagosome membrane expansion is mediated by the N-terminus and cis-membrane association of human ATG8s
eLife 12:e89185.
https://doi.org/10.7554/eLife.89185
