Assaying protein palmitoylation in plants

Plant Methods
Piers A HemsleyClaire S Grierson

Abstract

Protein S-acylation (also known as palmitoylation) is the reversible post-translational addition of acyl lipids to cysteine residues in proteins through a thioester bond. It allows strong association with membranes. Whilst prediction methods for S-acylation exist, prediction is imperfect. Existing protocols for demonstrating the S-acylation of plant proteins are either laborious and time consuming or expensive. We describe a biotin switch method for assaying the S-acylation of plant proteins. We demonstrate the technique by showing that the heterotrimeric G protein subunit AGG2 is S-acylated as predicted by mutagenesis experiments. We also show that a proportion of the Arabidopsis alpha-tubulin subunit pool is S-acylated in planta. This may account for the observed membrane association of plant microtubules. As alpha-tubulins are ubiquitously expressed they can potentially be used as a positive control for the S-acylation assay regardless of the cell type under study. We provide a robust biotin switch protocol that allows the rapid assay of protein S-acylation state in plants, using standard laboratory techniques and without the need for expensive or specialised equipment. We propose alpha-tubulin as a useful positive control f...Continue Reading

References

Sep 1, 1985·Analytical Biochemistry·B J NikolauP K Stumpf
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Jan 6, 2005·The Journal of Biological Chemistry·Eugenia G PolitisNicholas G Davis
Apr 23, 2005·Proceedings of the National Academy of Sciences of the United States of America·Han-Suk KimJeffery L Dangl
Aug 16, 2005·The Plant Cell·Piers A HemsleyClaire S Grierson
Nov 23, 2005·Proceedings of the National Academy of Sciences of the United States of America·Haitong HouChristian Ungermann
Jun 6, 2006·Cell·Amy F RothNicholas G Davis
Sep 12, 2006·Molecular & Cellular Proteomics : MCP·Frédéric FrottinThierry Meinnel
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Dec 13, 2006·Journal of Cell Science·Merel J W Adjobo-HermansTheodorus W J Gadella

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Citations

Feb 5, 2009·Molecular Membrane Biology·Piers A Hemsley
Oct 7, 2014·The New Phytologist·Piers A Hemsley
Dec 21, 2012·The New Phytologist·Piers A HemsleyClaire S Grierson
Feb 3, 2012·Cell Biology International·Maria KwiatkowskaJustyna Teresa Polit
Feb 20, 2015·Journal of Integrative Plant Biology·Zengyu LiuYi Zhang
Jun 14, 2013·Plant Signaling & Behavior·Piers A HemsleyClaire Grierson
Apr 3, 2020·Biochemical Society Transactions·Piers A Hemsley
Nov 7, 2019·Plant Biotechnology Journal·Xiaomin ZhangFeng Qin
Apr 11, 2017·Frontiers in Plant Science·Yaxiao Li, Baoxiu Qi
Feb 6, 2017·Journal of Experimental Botany·Piers A Hemsley

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Methods Mentioned

BETA
transgenic
immunoprecipitation
acylation
affinity

Software Mentioned

Palm
Terminator2
CSS
NBA

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