Abstract
Macroautophagy (hereafter referred to as autophagy) is a highly conserved catabolic eukaryotic pathway that is critical for stress responses and homeostasis. Atg18, one of the core proteins involved in autophagy, belongs to the PROPPIN family and is composed of seven WD40 repeats. Together with Atg2, Atg18 participates in the elongation of phagophores and the recycling of Atg9 in yeast. Despite extensive studies on the PROPPIN family, the structure of Atg18 from Saccharomyces cerevisiae has not been determined. Here, we report the structure of ScAtg18 at a resolution of 2.8 Å. Based on bioinformatics and structural analysis, we found that the 7AB loop of ScAtg18 is extended in Atg18, in comparison to other members of the PROPPIN family. Genetic analysis revealed that the 7AB loop of ScAtg18 is required for autophagy. Biochemical and biophysical experiments indicated that the 7AB loop of ScAtg18 is critical for interaction with ScAtg2 and the recruitment of ScAtg2 to the autophagy-initiating site. Collectively, our results show that the 7AB loop of ScAtg18 is a new binding site for Atg2 and is of functional importance to autophagy.
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Data availability
The coordinates and structural factors of the structure reported here have been deposited at the Protein Data Bank with PDB Code: 6KYB.
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Acknowledgements
We thank Jianhua He, Wenming Qin, Huan Zhou, and Feng Yu from BL17U, BL18U, and BL19U at the National Facility for Protein Science in Shanghai, Zhangjiang Laboratory (NFPS, ZJLab), China for providing technical support and assistance in data collection and analysis. This work was supported by National Key R&D Program of China grant 2017YFA0506300 (L. K.), NSFC grant 81671388 (Q. S.), NSFC grant 31770820 (L. K.), and NSFC grant 81772275 (G. L.).
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Q. S. and L. K. initiated the project. T. D., L. G., C.Q., X. L. and Q. S. performed the structural biological study. L.Y. and L.K. performed the cell biological experiments. Q. S. and L. K. wrote the manuscript. All the authors discussed the results and commented on the manuscript.
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Supplementary file1 Figure S1. Gel filtration profiles of ScAtg18 and ScAtg18∆433.The elution profiles of different proteins from Superdex 75 10/300 GL column are shown here. Red: ScAtg18; blue: ScAtg18∆433. The peak fraction of ScAtg18∆433 was visualized via the Coomassie blue-stained SDS-PAGE (PDF 665 kb)
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Lei, Y., Tang, D., Liao, G. et al. The crystal structure of Atg18 reveals a new binding site for Atg2 in Saccharomyces cerevisiae. Cell. Mol. Life Sci. 78, 2131–2143 (2021). https://doi.org/10.1007/s00018-020-03621-9
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DOI: https://doi.org/10.1007/s00018-020-03621-9