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An atlas of Arabidopsis protein S-acylation reveals its widespread role in plant cell organization and function

Nature Plants volume 8, pages 670–681 (2022)Cite this article

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Abstract

S-acylation is the addition of a fatty acid to a cysteine residue of a protein. While this modification may profoundly alter protein behaviour, its effects on the function of plant proteins remains poorly characterized, largely as a result of the lack of basic information regarding which proteins are S-acylated and where in the proteins the modification occurs. To address this gap in our knowledge, we used an optimized acyl-resin-assisted capture assay to perform a comprehensive analysis of plant protein S-acylation from six separate tissues. In our high- and medium-confidence groups, we identified 1,849 cysteines modified by S-acylation, which were located in 1,640 unique peptides from 1,094 different proteins. This represents around 6% of the detectable Arabidopsis proteome and suggests an important role for S-acylation in many essential cellular functions including trafficking, signalling and metabolism. To illustrate the potential of this dataset, we focus on cellulose synthesis and confirm the S-acylation of a number of proteins known to be involved in cellulose synthesis and trafficking of the cellulose synthase complex. In the secondary cell walls, cellulose synthesis requires three different catalytic subunits (CESA4, CESA7 and CESA8) that all exhibit striking sequence similarity and are all predicted to possess a RING-type zinc finger at their amino terminus composed of eight cysteines. For CESA8, we find evidence for S-acylation of these cysteines that is incompatible with any role in coordinating metal ions. We show that while CESA7 may possess a RING-type domain, the same region of CESA8 appears to have evolved a very different structure. Together, the data suggest that this study represents an atlas of S-acylation in Arabidopsis that will facilitate the broader study of this elusive post-translational modification in plants as well as demonstrating the importance of further work in this area.

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Fig. 1: Overview of the Arabidopsis acylome.
Fig. 2: Validation of S-acylated proteins using acyl-RAC assays and immunoblotting.
Fig. 3: S-acylation of KOR1.
Fig. 4: Differential S-acylation of CESA isoforms.
Fig. 5: Analysis of RING domain cysteines in CESA7, CESA4 and CESA8.

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Data availability

All processed data are included with the manuscript as supplementary tables. The unprocessed MS proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE63 partner repository with the dataset identifier PXD031348 and https://doi.org/10.6019/PXD031348. All figures have associated raw data, which will be provided upon request. Araport11 (https://www.arabidopsis.org/index.jsp) and E. coli UP000000625 were used as the protein databases for Arabidopsis and E. coli for performing MS database searches. Source data are provided with this paper.

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Acknowledgements

This work was supported by grants from the BBSRC (reference no. BB/P01013X/1) and Leverhulme Trust (no. RPG-2020-257). We acknowledge help provided by S. Warwood, J. Selley, R. O’cualain, D. Knight and E. Keevill at the Bio-MS core facility, RRID: SCR_020987, University of Manchester, for the proteomic analysis. We thank S. Persson for providing the cmu1 cmu2, cc1 cc2 and csi1 mutants, S. Vernhettes for providing the kor1-1 mutants, and T. Nuhse and O. Quinn for their comments on the manuscript.

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  1. Paul Carr

    Present address: Holiferm, Manchester, UK

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  1. Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK

    Manoj Kumar, Paul Carr & Simon R. Turner

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  1. Manoj Kumar
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Contributions

M.K. and S.T. conceived and planned the experiments. M.K. and P.C. carried out the experiments. M.K. and S.T. interpreted the results and wrote the manuscript.

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Correspondence to Simon R. Turner.

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Nature Plants thanks Herman Hofte, Ying Gu and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Supplementary Table 1.

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Kumar, M., Carr, P. & Turner, S.R. An atlas of Arabidopsis protein S-acylation reveals its widespread role in plant cell organization and function. Nat. Plants 8, 670–681 (2022). https://doi.org/10.1038/s41477-022-01164-4

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