Non-specific effects of the CINNAMATE-4-HYDROXYLASE inhibitor piperonylic acid

Ilias El Houari, Petr Klíma, Alexandra Baekelandt, Paul E. Staswick, Veselina Uzunova, Charo I. Del Genio, Ward Steenackers, Petre I. Dobrev, Roberta Filepová, Ondrej Novák, Richard Napier, Jan Petrášek, Dirk Inzé, Wout Boerjan, Bartel Vanholme

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1 Citation (Scopus)


Chemical inhibitors are often implemented for the functional characterization of genes to overcome the limitations associated with genetic approaches. Although it is well established that the specificity of the compound is key to success of a pharmacological approach, off-target effects are often overlooked or simply neglected in a complex biological setting. Here we illustrate the cause and implications of such secondary effects by focusing on piperonylic acid (PA), an inhibitor of CINNAMATE-4-HYDROXYLASE (C4H) that is frequently used to investigate the involvement of lignin during plant growth and development. When supplied to plants, we found that PA is recognized as a substrate by GRETCHEN HAGEN 3.6 (GH3.6), an amido synthetase involved in the formation of the indole-3-acetic acid (IAA) conjugate IAA-Asp. By competing for the same enzyme, PA interferes with IAA conjugation, resulting in an increase in IAA concentrations in the plant. In line with the broad substrate specificity of the GH3 family of enzymes, treatment with PA increased not only IAA levels but also those of other GH3-conjugated phytohormones, namely jasmonic acid and salicylic acid. Finally, we found that interference with the endogenous function of GH3s potentially contributes to phenotypes previously observed upon PA treatment. We conclude that deregulation of phytohormone homeostasis by surrogate occupation of the conjugation machinery in the plant is likely a general phenomenon when using chemical inhibitors. Our results hereby provide a novel and important basis for future reference in studies using chemical inhibitors.

Original languageEnglish
Pages (from-to)470-479
Number of pages10
JournalThe Plant Journal
Issue number2
Early online date10 Apr 2023
Publication statusPublished - Jul 2023

Bibliographical note

This is the peer reviewed version of the following article: El Houari, I, Klíma, P, Baekelandt, A, Staswick, PE, Uzunova, V, Del Genio, CI, Steenackers, W, Dobrev, PI, Filepová, R, Novák, O, Napier, R, Petrášek, J, Inzé, D, Boerjan, W & Vanholme, B 2023, 'Non-specific effects of the CINNAMATE-4-HYDROXYLASE inhibitor piperonylic acid', The Plant Journal, vol. 115, no. 2, pp. 470-479, which has been published in final form at This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. This article may not be enhanced, enriched or otherwise transformed into a derivative work, without express permission from Wiley or by statutory rights under applicable legislation. Copyright notices must not be removed, obscured or modified. The article must be linked to Wiley’s version of record on Wiley Online Library and any embedding, framing or otherwise making available the article or pages thereof by third parties from platforms, services and websites other than Wiley Online Library must be prohibited.

This document is the author’s post-print version, incorporating any revisions agreed during the peer-review process. Some differences between the published version and this version may remain and you are advised to consult the published version if you wish to cite from it.


Belgian American Educational Foundation
European Molecular Biology Organization. Grant Number: STF-8658
Fonds Wetenschappelijk Onderzoek. Grant Numbers: 1S04020N, 3G038719, G008116N, V414521N
Ministry of Education, Youth, and Sports of the Czech Republic. Grant Number: CZ.02.1.01/0.0/0.0/16_019/0000738
U.S. Department of Agriculture
UK Research and Innovation. Grant Number: MD/T020652/1


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