Alterations of hydrogen peroxide (H2O2) levels have a profound impact on numerous signaling cascades orchestrating plant growth, development, and stress signaling, including programmed cell death. To expand the repertoire of known molecular mechanisms implicated in H2O2 signaling, we performed a forward chemical screen to identify small molecules that could alleviate the photorespiratory-induced cell death phenotype of Arabidopsisthaliana mutants lacking H2O2-scavenging capacity by peroxisomal catalase2. Here, we report the characterization of pakerine, an m-sulfamoyl benzamide from the sulfonamide family. Pakerine alleviates the cell death phenotype of cat2 mutants exposed to photorespiration-promoting conditions and delays dark-induced senescence in wild-type Arabidopsis leaves. By using a combination of transcriptomics, metabolomics, and affinity purification, we identified abnormal inflorescence meristem 1 (AIM1) as a putative protein target of pakerine. AIM1 is a 3-hydroxyacyl-CoA dehydrogenase involved in fatty acid β-oxidation that contributes to jasmonic acid (JA) and salicylic acid (SA) biosynthesis. Whereas intact JA biosynthesis was not required for pakerine bioactivity, our results point toward a role for β-oxidation-dependent SA production in the execution of H2O2-mediated cell death.

Department of Biomolecular Medicine Faculty of Medicine and Health Sciences Ghent University B 9000 Ghent Belgium

Department of Green Chemistry and Technology Faculty of Bioscience Engineering Ghent University B 9000 Ghent Belgium

Department of Molecular Biology and Radiobiology Faculty of AgriSciences Mendel University in Brno 61300 Brno Czech Republic

Department of Plant Biotechnology and Bioinformatics Ghent University B 9052 Ghent Belgium

Laboratory of Plant Physiology Plant Sciences Group Wageningen University and Research 6708 PB Wageningen The Netherlands

Phytophthora Research Centre Mendel University in Brno 61300 Brno Czech Republic

VIB Center for Medical Biotechnology B 9052 Ghent Belgium

VIB Center for Plant Systems Biology B 9052 Ghent Belgium

VIB Proteomics Core B 9000 Ghent Belgium

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Mhamdi A., Van Breusegem F. Reactive oxygen species in plant development. Development. 2018;145:dev164376. doi: 10.1242/dev.164376. PubMed DOI

Choudhury F.K., Rivero R.M., Blumwald E., Mittler R. Reactive oxygen species, abiotic stress and stress combination. Plant J. 2017;90:856–867. doi: 10.1111/tpj.13299. PubMed DOI

Noctor G., Reichheld J.P., Foyer C.H. ROS-related redox regulation and signaling in plants. Semin. Cell Dev. Biol. 2018;80:3–12. doi: 10.1016/j.semcdb.2017.07.013. PubMed DOI

Smirnoff N., Arnaud D. Hydrogen peroxide metabolism and functions in plants. New Phytol. 2019;221:1197–1214. doi: 10.1111/nph.15488. PubMed DOI

Levine A., Tenhaken R., Dixon R., Lamb C. H2O2 from the oxidative burst orchestrates the plant hypersensitive disease resistance response. Cell. 1994;79:583–593. doi: 10.1016/0092-8674(94)90544-4. PubMed DOI

Willems P., Mhamdi A., Stael S., Storme V., Kerchev P., Noctor G., Gevaert K., Van Breusegem F. The ROS wheel: Refining ROS transcriptional footprints. Plant Physiol. 2016;171:1720–1733. doi: 10.1104/pp.16.00420. PubMed DOI PMC

Vanderauwera S., Zimmermann P., Rombauts S., Vandenabeele S., Langebartels C., Gruissem W., Inzé D., Van Breusegem F. Genome-wide analysis of hydrogen peroxide-regulated gene expression in Arabidopsis reveals a high light-induced transcriptional cluster involved in anthocyanin biosynthesis. Plant Physiol. 2005;139:806–821. doi: 10.1104/pp.105.065896. PubMed DOI PMC

Huang J., Willems P., Van Breusegem F., Messens J. Pathways crossing mammalian and plant sulfenomic landscapes. Free Radic. Biol. Med. 2018;122:193–201. doi: 10.1016/j.freeradbiomed.2018.02.012. PubMed DOI

Huang J., Willems P., Wei B., Tian C., Ferreira R.B., Bodra N., Martínez Gache S.A., Wahni K., Liu K., Vertommen D., et al. Mining for protein S-sulfenylation in Arabidopsis uncovers redox-sensitive sites. Proc. Natl. Acad. Sci. USA. 2019;116:20256–20261. doi: 10.1073/pnas.1906768116. PubMed DOI PMC

Motohashi K., Koyama F., Nakanishi Y., Ueoka-Nakanishi H., Hisabori T. Chloroplast Cyclophilin Is a Target Protein of Thioredoxin. Thiol modulation of the peptidyl-prolyl cis-trans isomerase activity. J. Biol. Chem. 2003;278:31848–31852. doi: 10.1074/jbc.M304258200. PubMed DOI

Park S.W., Li W., Viehhauser A., He B., Kim S., Nilsson A.K., Andersson M.X., Kittle J.D., Ambavaram M.M.R., Luan S., et al. Cyclophilin 20-3 relays a 12-oxo-phytodienoic acid signal during stress responsive regulation of cellular redox homeostasis. Proc. Natl. Acad. Sci. USA. 2013;110:9559–9564. doi: 10.1073/pnas.1218872110. PubMed DOI PMC

Tian Y., Fan M., Qin Z., Lv H., Wang M., Zhang Z., Zhou W., Zhao N., Li X., Han C., et al. Hydrogen peroxide positively regulates brassinosteroid signaling through oxidation of the BRASSINAZOLE-RESISTANT1 transcription factor. Nat. Commun. 2018;9:1063. doi: 10.1038/s41467-018-03463-x. PubMed DOI PMC

Tuzet A., Rahantaniaina M.S., Noctor G. Analyzing the Function of Catalase and the Ascorbate-Glutathione Pathway in H2O2 Processing: Insights from an Experimentally Constrained Kinetic Model. Antioxid. Redox Signal. 2019;30:1238–1268. doi: 10.1089/ars.2018.7601. PubMed DOI

Timm S., Hagemann M. Photorespiration—How is it regulated and regulates overall plant metabolism. J. Exp. Bot. 2020;71:3955–3965. doi: 10.1093/jxb/eraa183. PubMed DOI

Queval G., Issakidis-Bourguet E., Hoeberichts F.A., Vandorpe M., Gakière B., Vanacker H., Miginiac-Maslow M., Van Breusegem F., Noctor G. Conditional oxidative stress responses in the Arabidopsis photorespiratory mutant cat2 demonstrate that redox state is a key modulator of daylength-dependent gene expression, and define photoperiod as a crucial factor in the regulation of H2O2-induced cel. Plant J. 2007;52:640–657. doi: 10.1111/j.1365-313X.2007.03263.x. PubMed DOI

Vandenabeele S., Van Der Kelen K., Dat J., Gadjev I., Boonefaes T., Morsa S., Rottiers P., Slooten L., Van Montagu M., Zabeau M., et al. A comprehensive analysis of hydrogen peroxide-induced gene expression in tobacco. Proc. Natl. Acad. Sci. USA. 2003;100:16113–16118. doi: 10.1073/pnas.2136610100. PubMed DOI PMC

Mhamdi A., Noctor G., Baker A. Plant catalases: Peroxisomal redox guardians. Arch. Biochem. Biophys. 2012;525:181–194. doi: 10.1016/j.abb.2012.04.015. PubMed DOI

Han Y., Mhamdi A., Chaouch S., Noctor G. Regulation of basal and oxidative stress-triggered jasmonic acid-related gene expression by glutathione. Plant. Cell Environ. 2013;36:1135–1146. doi: 10.1111/pce.12048. PubMed DOI

Chaouch S., Queval G., Vanderauwera S., Mhamdi A., Vandorpe M., Langlois-Meurinne M., Van Breusegem F., Saindrenan P., Noctor G. Peroxisomal hydrogen peroxide is coupled to biotic defense responses by ISOCHORISMATE SYNTHASE1 in a daylength-related manner. Plant Physiol. 2010;153:1692–1705. doi: 10.1104/pp.110.153957. PubMed DOI PMC

Han Y., Chaouch S., Mhamdi A., Queval G., Zechmann B., Noctor G. Functional Analysis of Arabidopsis Mutants Points to Novel Roles for Glutathione in Coupling H2O2 to Activation of Salicylic Acid Accumulation and Signaling. Antioxid. Redox Signal. 2013;18:2106–2121. doi: 10.1089/ars.2012.5052. PubMed DOI PMC

Mhamdi A., Mauve C., Houda G., Saindrenan P., Hodges M., Noctor G. Cytosolic NADP-dependent isocitrate dehydrogenase contributes to redox homeostasis and the regulation of pathogen responses in Arabidopsis leaves. Plant. Cell Environ. 2010;33:1112–1123. doi: 10.1111/j.1365-3040.2010.02133.x. PubMed DOI

Kaurilind E., Xu E., Brosché M. A genetic framework for H2O2 induced cell death in Arabidopsis thaliana. BMC Genom. 2015;16:837. doi: 10.1186/s12864-015-1964-8. PubMed DOI PMC

Kerchev P., Waszczak C., Lewandowska A., Willems P., Shapiguzov A., Li Z., Alseekh S., Mühlenbock P., Hoeberichts F.A., Huang J., et al. Lack of GLYCOLATE OXIDASE1, but Not GLYCOLATE OXIDASE2, Attenuates the Photorespiratory Phenotype of CATALASE2-Deficient Arabidopsis. Plant Physiol. 2016;171:1704–1719. doi: 10.1104/pp.16.00359. PubMed DOI PMC

Waszczak C., Kerchev P.I., Mühlenbock P., Hoeberichts F.A., Van Der Kelen K., Mhamdi A., Willems P., Denecker J., Kumpf R.P., Noctor G., et al. SHORT-ROOT Deficiency Alleviates the Cell Death Phenotype of the Arabidopsis catalase2 Mutant under Photorespiration-Promoting Conditions. Plant Cell. 2016;28:1844–1859. doi: 10.1105/tpc.16.00038. PubMed DOI PMC

Meinke D., Muralla R., Sweeney C., Dickerman A. Identifying essential genes in Arabidopsis thaliana. Trends Plant Sci. 2008;13:483–491. doi: 10.1016/j.tplants.2008.06.003. PubMed DOI

Park S.-Y., Fung P., Nishimura N., Jensen D.R., Fujii H., Zhao Y., Lumba S., Santiago J., Rodrigues A., Chow T.-F.F., et al. Abscisic acid inhibits type 2C protein phosphatases via the PYR/PYL family of START proteins. Science. 2009;324:1068–1071. doi: 10.1126/science.1173041. PubMed DOI PMC

De Rybel B., Audenaert D., Vert G., Rozhon W., Mayerhofer J., Peelman F., Coutuer S., Denayer T., Jansen L., Nguyen L., et al. Chemical Inhibition of a Subset of Arabidopsis thaliana GSK3-like Kinases Activates Brassinosteroid Signaling. Chem. Biol. 2009;16:594–604. doi: 10.1016/j.chembiol.2009.04.008. PubMed DOI PMC

Kerchev P., van der Meer T., Sujeeth N., Verlee A., Stevens C.V., Van Breusegem F., Gechev T. Molecular priming as an approach to induce tolerance against abiotic and oxidative stresses in crop plants. Biotechnol. Adv. 2020;40:107503. doi: 10.1016/j.biotechadv.2019.107503. PubMed DOI

Bussell J.D., Reichelt M., Wiszniewski A.A.G., Gershenzon J., Smith S.M. Peroxisomal ATP-binding cassette transporter COMATOSE and the multifunctional protein abnormal INFLORESCENCE MERISTEM are required for the production of benzoylated metabolites in Arabidopsis seeds. Plant Physiol. 2014;164:48–54. doi: 10.1104/pp.113.229807. PubMed DOI PMC

Chamnongpol S., Willekens H., Moeder W., Langebartels C., Sandermann H., Van Montagu M., Inzé D., Van Camp W. Defense activation and enhanced pathogen tolerance induced by H2O2 in transgenic tobacco. Proc. Natl. Acad. Sci. USA. 1998;95:5818–5823. doi: 10.1073/pnas.95.10.5818. PubMed DOI PMC

May M.J., Leaver C.J. Oxidative Stimulation of Glutathione Synthesis in Arabidopsis thaliana Suspension Cultures. Plant Physiol. 1993;103:621–627. doi: 10.1104/pp.103.2.621. PubMed DOI PMC

Meijering E., Jacob M., Sarria J.-C.F., Steiner P., Hirling H., Unser M. Design and validation of a tool for neurite tracing and analysis in fluorescence microscopy images. Cytometry. 2004;58:167–176. doi: 10.1002/cyto.a.20022. PubMed DOI

Robinson M., Mccarthy D., Chen Y., Smyth G.K. edgeR: Differential expression analysis of digital gene expression data. Bioinformatics. 2010;26:139–140. doi: 10.1093/bioinformatics/btp616. PubMed DOI PMC

Verlee A., Heugebaert T., Van Der Meer T., Kerchev P.I., Van Breusegem F., Stevens C.V. A chemoselective and continuous synthesis of m-sulfamoylbenzamide analogues. Beilstein J. Org. Chem. 2017;13:303–312. doi: 10.3762/bjoc.13.33. PubMed DOI PMC

Kerchev P., Mühlenbock P., Denecker J., Morreel K., Hoeberichts F., Van der Kelen K., Vandorpe M., Nguyen L., Audenaert D., Van Breusegem F. Activation of auxin signalling counteracts photorespiratory H 2 O 2 -dependent cell death. Plant Cell Environ. 2015;38:253–265. doi: 10.1111/pce.12250. PubMed DOI

Vanderauwera S., Vandenbroucke K., Inzé A., Van De Cotte B., Mühlenbock P., De Rycke R., Naouar N., Van Gaever T., Van Montagu M.C.E., Van Breusegem F. AtWRKY15 perturbation abolishes the mitochondrial stress response that steers osmotic stress tolerance in Arabidopsis. Proc. Natl. Acad. Sci. USA. 2012;109:20113–20118. doi: 10.1073/pnas.1217516109. PubMed DOI PMC

Hruz T., Laule O., Szabo G., Wessendorp F., Bleuler S., Oertle L., Widmayer P., Gruissem W., Zimmermann P. Resource Review Genevestigator V3: A Reference Expression Database for the Meta-Analysis of Transcriptomes. Adv. Bioinform. 2008 doi: 10.1155/2008/420747. PubMed DOI PMC

Van Doorn W.G., Woltering E.J. Senescence and programmed cell death: Substance or semantics? J. Exp. Bot. 2004;55:2147–2153. doi: 10.1093/jxb/erh264. PubMed DOI

Weaver L.M., Amasino R.M. Senescence is induced in individually darkened Arabidopsis leaves, but inhibited in whole darkened plants. Plant Physiol. 2001;127:876–886. doi: 10.1104/pp.010312. PubMed DOI PMC

Jenmalm Jensen A., Cornella Taracido I. Affinity-Based Chemoproteomics for Target Identification. In: Plowright A.T., editor. Target Discovery and Validation: Methods and Strategies for Drug Discovery. Wiley-VCH Verlag GmbH & Co. KGaA; Weinheim, Germany: 2019. pp. 25–49. DOI

Le B.H., Cheng C., Bui A.Q., Wagmaister J.A., Henry K.F., Pelletier J., Kwong L., Belmonte M., Kirkbride R., Horvath S., et al. Global analysis of gene activity during Arabidopsis seed development and identification of seed-specific transcription factors. Proc. Natl. Acad. Sci. USA. 2010;107:8063–8070. doi: 10.1073/pnas.1003530107. PubMed DOI PMC

Kombrink E. Chemical and genetic exploration of jasmonate biosynthesis and signaling paths. Planta. 2012;236:1351–1366. doi: 10.1007/s00425-012-1705-z. PubMed DOI

Delker C., Zolman B.K., Miersch O., Wasternack C. Jasmonate biosynthesis in Arabidopsis thaliana requires peroxisomal β-oxidation enzymes—Additional proof by properties of pex6 and aim1. Phytochemistry. 2007;68:1642–1650. doi: 10.1016/j.phytochem.2007.04.024. PubMed DOI

García-Galán M.J., Silvia Díaz-Cruz M., Barceló D. Identification and determination of metabolites and degradation products of sulfonamide antibiotics. TrAC Trends Anal. Chem. 2008;27:1008–1022. doi: 10.1016/j.trac.2008.10.001. DOI

Brazier-Hicks M., Howell A., Cohn J., Hawkes T., Hall G., Mcindoe E., Edwards R. Chemically induced herbicide tolerance in rice by the safener metcamifen is associated with a phased stress response. J. Exp. Bot. 2020;71:411–421. doi: 10.1093/jxb/erz438. PubMed DOI PMC

Giannakopoulos G., Dittgen J., Schulte W., Zoellner P., Helmke H., Lagojda A., Edwards R. Safening activity and metabolism of the safener cyprosulfamide in maize and wheat. Pest Manag. Sci. 2020:ps.5801. doi: 10.1002/ps.5801. PubMed DOI

Okamoto M., Peterson F.C., Defries A., Park S.-Y., Endo A., Nambara E., Volkman B.F., Cutler S.R. Activation of dimeric ABA receptors elicits guard cell closure, ABA-regulated gene expression, and drought tolerance. Proc. Natl. Acad. Sci. USA. 2013;110:12132–12137. doi: 10.1073/pnas.1305919110. PubMed DOI PMC

Cao M.-J., Zhang Y.-L., Liu X., Huang H., Zhou X.E., Wang W.-L., Zeng A., Zhao C.-Z., Si T., Du J., et al. Combining chemical and genetic approaches to increase drought resistance in plants. Nat. Commun. 2017;8:1183. doi: 10.1038/s41467-017-01239-3. PubMed DOI PMC

Calabrese E.J., Mattson M.P. How does hormesis impact biology, toxicology, and medicine? NPJ Aging Mech. Dis. 2017;3:13. doi: 10.1038/s41514-017-0013-z. PubMed DOI PMC

Arent S., Christensen C.E., Pye V.E., Nørgaard A., Henriksen A. The multifunctional protein in peroxisomal beta-oxidation: Structure and substrate specificity of the Arabidopsis thaliana protein MFP2. J. Biol. Chem. 2010;285:24066–24077. doi: 10.1074/jbc.M110.106005. PubMed DOI PMC

Richmond T.A., Bleecker A.B. A Defect in-Oxidation Causes Abnormal Inflorescence Development in Arabidopsis. Plant Cell. 1999;11:1911–1924. doi: 10.1105/tpc.11.10.1911. PubMed DOI PMC

Rylott E.L., Eastmond P.J., Gilday A.D., Slocombe S.P., Larson T.R., Baker A., Graham I.A. The Arabidopsis thaliana multifunctional protein gene (MFP2) of peroxisomal β-oxidation is essential for seedling establishment. Plant J. 2006;45:930–941. doi: 10.1111/j.1365-313X.2005.02650.x. PubMed DOI

Zolman B.K., Yoder A., Bartel B. Genetic Analysis of Indole-3-butyric Acid Responses in Arabidopsis thaliana Reveals Four Mutant Classes. Genetics. 2000;156:1323–1337. PubMed PMC

Li Y., Liu Y., Zolman B.K. Metabolic alterations in the enoyl-coA hydratase 2 mutant disrupt peroxisomal pathways in seedlings. Plant Physiol. 2019;180:1860–1876. doi: 10.1104/pp.19.00300. PubMed DOI PMC

Troncoso-Ponce M.A., Cao X., Yang Z., Ohlrogge J.B. Lipid turnover during senescence. Plant Sci. 2013;205–206:13–19. doi: 10.1016/j.plantsci.2013.01.004. PubMed DOI

Castillo M.C., León J. Expression of the β-oxidation gene 3-ketoacyl-CoA thiolase 2 (KAT2) is required for the timely onset of natural and dark-induced leaf senescence in Arabidopsis. J. Exp. Bot. 2008;59:2171–2179. doi: 10.1093/jxb/ern079. PubMed DOI PMC

Kunz H.H., Scharnewski M., Feussner K., Feussner I., Flügge U.I., Fulda M., Gierth M. The ABC transporter PXA1 and peroxisomal β-oxidation are vital for metabolism in mature leaves of Arabidopsis during extended darkness. Plant Cell. 2009;21:2733–2749. doi: 10.1105/tpc.108.064857. PubMed DOI PMC

Knight Z.A., Shokat K.M. Chemical Genetics: Where Genetics and Pharmacology Meet. Cell. 2007;128:425–430. doi: 10.1016/j.cell.2007.01.021. PubMed DOI

Van Der Graaff E., Schwacke R., Schneider A., Desimone M., Flügge U.I., Kunze R. Transcription analysis of arabidopsis membrane transporters and hormone pathways during developmental and induced leaf senescence. Plant Physiol. 2006;141:776–792. doi: 10.1104/pp.106.079293. PubMed DOI PMC

Quirino B.F., Normanly J., Amasino R.M. Diverse range of gene activity during Arabidopsis thaliana leaf senescence includes pathogen-independent induction of defense-related genes. Plant Mol. Biol. 1999;40:267–278. doi: 10.1023/A:1006199932265. PubMed DOI

Kao Y.T., Gonzalez K.L., Bartel B. Peroxisome function, biogenesis, and dynamics in plants. Plant Physiol. 2018;176:162–177. doi: 10.1104/pp.17.01050. PubMed DOI PMC

Sequera-Mutiozabal M.I., Erban A., Kopka J., Atanasov K.E., Bastida J., Fotopoulos V., Alcázar R., Tiburcio A.F. Global metabolic profiling of arabidopsis polyamine oxidase 4 (AtPAO4) loss-of-function mutants exhibiting delayed dark-induced senescence. Front. Plant Sci. 2016;7:173. doi: 10.3389/fpls.2016.00173. PubMed DOI PMC

Chen D., Shao Q., Yin L., Younis A., Zheng B. Polyamine function in plants: Metabolism, regulation on development, and roles in abiotic stress responses. Front. Plant Sci. 2019;9:1945. doi: 10.3389/fpls.2018.01945. PubMed DOI PMC

Yang J., Duan G., Li C., Liu L., Han G., Zhang Y., Wang C. The Crosstalks Between Jasmonic Acid and Other Plant Hormone Signaling Highlight the Involvement of Jasmonic Acid as a Core Component in Plant Response to Biotic and Abiotic Stresses. Front. Plant Sci. 2019;10:1349. doi: 10.3389/fpls.2019.01349. PubMed DOI PMC

Balfagón D., Sengupta S., Gómez-Cadenas A., Fritschi F.B., Azad R.K., Mittler R., Zandalinasc S.I. Jasmonic acid is required for plant acclimation to a combination of high light and heat stress. Plant Physiol. 2019;181:1668–1682. doi: 10.1104/pp.19.00956. PubMed DOI PMC

Zhao Y., Dong W., Zhang N., Ai X., Wang M., Huang Z., Xiao L., Xia G. A wheat allene oxide cyclase gene enhances salinity tolerance via jasmonate signaling. Plant Physiol. 2014;164:1068–1076. doi: 10.1104/pp.113.227595. PubMed DOI PMC

Xu L., Zhao H., Ruan W., Deng M., Wang F., Peng J., Luo J., Chen Z., Yi K. ABNORMAL INFLORESCENCE MERISTEM1 functions in salicylic acid biosynthesis to maintain proper reactive oxygen species levels for root meristem activity in rice. Plant Cell. 2017;29:560–574. doi: 10.1105/tpc.16.00665. PubMed DOI PMC

Lefevere H., Bauters L., Gheysen G. Salicylic Acid Biosynthesis in Plants. Front. Plant Sci. 2020;11:338. doi: 10.3389/fpls.2020.00338. PubMed DOI PMC

Pallas J.A., Paiva N.L., Lamb C., Dixon R.A. Tobacco plants epigenetically suppressed in phenylalanine ammonia-lyase expression do not develop systemic acquired resistance in response to infection by tobacco mosaic virus. Plant J. 1996;10:281–293. doi: 10.1046/j.1365-313X.1996.10020281.x. DOI

Shine M.B., Yang J.-W., El-Habbak M., Nagyabhyru P., Fu D.-Q., Navarre D., Ghabrial S., Kachroo P., Kachroo A. Cooperative functioning between phenylalanine ammonia lyase and isochorismate synthase activities contributes to salicylic acid biosynthesis in soybean. New Phytol. 2016;212:627–636. doi: 10.1111/nph.14078. PubMed DOI