N-Sulfonated IAA was discovered as a novel auxin metabolite in Urtica where it is biosynthesized de novo utilizing inorganic sulfate. It showed no auxin activity in DR5::GUS assay, implying possible inactivation/storage mechanism. A novel auxin derivative, N-sulfoindole-3-acetic acid (IAA-N-SO3H, SIAA), was discovered in stinging nettle (Urtica dioica) among 116 sulfonated metabolites putatively identified by a semi-targeted UHPLC-QqTOF-MS analysis of 23 plant/algae/fungi species. These sulfometabolites were detected based on the presence of a neutral loss of sulfur trioxide, as indicated by the m/z difference of 79.9568 Da in the MS2 spectra. The structure of newly discovered SIAA was confirmed by synthesizing its standard and comparing retention time, m/z and MS2 spectrum with those of SIAA found in Urtica. To study its natural occurrence, 73 species in total were further analyzed by UHPLC-QqTOF-MS or targeted UHPLC-MS/MS method with a limit of detection of 244 fmol/g dry weight. However, SIAA was only detected in Urtica at a concentration of 13.906 ± 9.603 nmol/g dry weight. Its concentration was > 30 times higher than that of indole-3-acetic acid (IAA), and the SIAA/IAA ratio was further increased under different light conditions, especially in continuous blue light. In addition to SIAA, structurally similar metabolites, N-sulfoindole-3-lactic acid, 4-(sulfooxy)phenyllactic acid and 4-(sulfooxy)phenylacetic acid, were detected in Urtica for the first time. SIAA was biosynthesized from inorganic sulfate in seedlings, as confirmed by the incorporation of exogenous 34S-ammonium sulfate (1 mM and 10 mM). SIAA exhibited no auxin activity, as demonstrated by both the Arabidopsis DR5::GUS assay and the Arabidopsis phenotype analysis. Sulfonation of IAA may therefore be a mechanism for IAA deactivation and/or storage in Urtica, similar to sulfonation of the jasmonates in Arabidopsis.

Czech Advanced Technology and Research Institute Palacký University Olomouc Šlechtitelů 27 CZ 77900 Olomouc Czech Republic

Department of Chemical Biology Palacký University Olomouc Šlechtitelů 27 CZ 77900 Olomouc Czech Republic

Department of Experimental Biology Palacký University Olomouc Šlechtitelů 27 CZ 77900 Olomouc Czech Republic

Laboratory of Growth Regulators Institute of Experimental Botany The Czech Academy of Sciences and Faculty of Science Palacký University Olomouc Czech Republic

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Abdullah Y, Schneider B, Petersen M (2008) Occurrence of rosmarinic acid, chlorogenic acid and rutin in DOI

Arcos M, Olivera ER, Arias S, Naharro G, Luengo JM (2010) The 3,4-dihydroxyphenylacetic acid catabolon, a catabolic unit for degradation of biogenic amines tyramine and dopamine in PubMed DOI

Baek D, Pathange P, Chung JS, Jiang JF, Gao LQ, Oikawa A, Hirai MY, Saito K, Pare PW, Shi HZ (2010) A stress-inducible sulphotransferase sulphonates salicylic acid and confers pathogen resistance in PubMed DOI

Bialek K, Cohen JD (1986) Isolation and partial characterization of the major amide-linked conjugate of indole-3-acetic-acid from phaseolus-vulgarIS L. Plant Physiol 80:99–104 PubMed DOI PMC

Boerjan W, Cervera MT, Delarue M, Beeckman T, Dewitte W, Bellini C, Caboche M, Vanonckelen H, Vanmontagu M, Inze D (1995) Superroot, a recessive mutation in arabidopsis, confers auxin overproduction. Plant Cell 7:1405–1419 PubMed PMC

Brunoni F, Pencík A, Zukauskaite A, Ament A, Kopecná M, Collani S, Kopecny D, Novák O (2023) Amino acid conjugation of oxIAA is a secondary metabolic regulation involved in auxin homeostasis. New Phytol 238:2264–2270 PubMed DOI

Cheynier V, Comte G, Davies KM, Lattanzio V, Martens S (2013) Plant phenolics: Recent advances on their biosynthesis, genetics, and ecophysiology. Plant Physiol Biochem 72:1–20 PubMed DOI

Davies PJ (2004) The plant hormones: Their nature, occurrence, and functions. Plant Hormones: Biosynthesis, Signal Transduction, Action. 1–15

De Diego N, Fürst T, Humplík JF, Ugena L, Podlesáková K, Spíchal L (2017) An automated method for high-throughput screening of PubMed DOI PMC

Di Q, Li Y, Zhang DP, Wu W, Zhang L, Zhao X, Luo L, Yu LL (2022) A novel type of phytosulfokine, PSK-ε, positively regulates root elongation and formation of lateral roots and root nodules in PubMed DOI PMC

Elliott MC, Stowe BB (1970) A novel sulphonated natural indole. Phytochemistry 9:1629–2000 DOI

Faulkner IJ, Rubery PH (1992) Flavonoids and flavonoid sulfates as probes of auxin-transport regulation in cucurbita-pepo hypocotyl segments and vesicles. Planta 186:618–625 PubMed DOI

Fernández-Milmanda GL, Crocco CD, Reichelt M, Mazza CA, Köllner TG, Zhang T, Cargnel MD, Lichy MZ, Fiorucci AS, Fankhauser C, Koo AJ, Austin AT, Gershenzon J, Ballaré CL (2020) A light-dependent molecular link between competition cues and defence responses in plants. Nature Plants 6:223–230 PubMed DOI

Gidda SK, Miersch O, Levitin A, Schmidt J, Wasternack C, Varin L (2003) Biochemical and molecular characterization of a hydroxyjasmonate sulfotransferase from Arabidopsis thaliana. J Biol Chem 278:17895–17900 PubMed DOI

Gläser K, Kanawati B, Kubo T, Schmitt-Kopplin P, Grill E (2014) Exploring the PubMed DOI

Günal S, Hardman R, Kopriva S, Mueller JW (2019) Sulfation pathways from red to green. J Biol Chem 294:12293–12312 PubMed DOI PMC

Halliday KJ, Fankhauser C (2003) Phytochrome-hormonal signalling networks. New Phytol 157:449–463 PubMed DOI

Hayashi K, Arai K, Aoi Y, Tanaka Y, Hira H, Guo RP, Hu Y, Ge CN, Zhao YD, Kasahara H, Fukui K (2021) The main oxidative inactivation pathway of the plant hormone auxin. Nat Commun 12:6752 PubMed DOI PMC

Hilbert M, Voll LM, Ding Y, Hofmann J, Sharma M, Zuccaro A (2012) Indole derivative production by the root endophyte PubMed DOI

Hirschmann F, Krause F, Papenbrock J (2014) The multi-protein family of sulfotransferases in plants: composition, occurrence, substrate specificity, and functions. Front Plant Sci. 10.3389/fpls.2014.00556 PubMed DOI PMC

Horng AJ, Yang SF (1975) Aerobic oxidation of indole-3-acetic-acid with bisulfite. Phytochemistry 14:1425–1428 DOI

Kai K, Wakasa K, Miyagawa H (2007) Metabolism of indole-3-acetic acid in rice: Identification and characterization of N-β-D-glucopyranosyl indole-3-acetic acid and its conjugates. Phytochemistry 68:2512–2522 PubMed DOI

Keuskamp DH, Sasidharan R, Pierik R (2010) Physiological regulation and functional significance of shade avoidance responses to neighbors. Plant Signal Behav 5:655–662 PubMed DOI PMC

Kleinenkuhnen N, Büchel F, Gerlich SC, Kopriva S, Metzger S (2019) A novel method for identification and quantification of sulfated flavonoids in plants by neutral loss scan mass spectrometry. Front Plant Sci. 10.3389/fpls.2019.00885 PubMed DOI PMC

Koprivova A, Kopriva S (2016) Sulfation pathways in plants. Chem Biol Interact 259:23–30 PubMed DOI

Korasick DA, Enders TA, Strader LC (2013) Auxin biosynthesis and storage forms. J Exp Bot 64:2541–2555 PubMed DOI PMC

Kurth C, Welling M, Pohnert G (2015) Sulfated phenolic acids from PubMed DOI

Lacomme C, Roby D (1996) Molecular cloning of a sulfotransferase in PubMed DOI

Lau OL, John WW, Yang SF (1978) Inactivity of oxidation-products of indole-3-acetic-acid on ethylene production in mung bean hypocotyls. Plant Physiol 61:68–71 PubMed DOI PMC

Li Y, Di Q, Luo L, Yu LL (2024) Phytosulfokine peptides, their receptors, and functions. Front Plant Sci. 10.3389/fpls.2023.1326964 PubMed DOI PMC

Ljung K (2013) Auxin metabolism and homeostasis during plant development. Development 140:943–950 PubMed DOI

Ludwig-Muller J (2011) Auxin conjugates: their role for plant development and in the evolution of land plants. J Exp Bot 62:1757–1773 PubMed DOI

Marsolais F, Boyd J, Paredes Y, Schinas AM, Garcia M, Elzein S, Varin L (2007) Molecular and biochemical characterization of two brassinosteroid sulfotransferases from PubMed DOI

Matsubayashi Y, Sakagami Y (1996) Phytosulfokine, sulfated peptides that induce the proliferation of single mesophyll cells of PubMed DOI PMC

Miersch O, Neumerkel J, Dippe M, Stenzel I, Wasternack C (2008) Hydroxylated jasmonates are commonly occurring metabolites of jasmonic acid and contribute to a partial switch-off in jasmonate signaling. New Phytol 177:114–127 PubMed DOI

Noleto-Dias C, Harflett C, Beale MH, Ward JL (2020) Sulfated flavanones and dihydroflavonols from willow. Phytochem Lett 35:88–93 PubMed DOI PMC

Nzowa LK, Teponno RB, Tapondjou LA, Verotta L, Liao Z, Graham D, Zink MC, Barboni L (2013) Two new tryptophan derivatives from the seed kernels of PubMed DOI PMC

Pencík A, Casanova-Sáez R, Pilarová V, Zukauskaite A, Pinto R, Micol JL, Ljung K, Novák O (2018) Ultra-rapid auxin metabolite profiling for high-throughput mutant screening in PubMed DOI PMC

Porco S, Pencík A, Rashed A, Voss U, Casanova-Sáez R, Bishopp A, Golebiowska A, Bhosale R, Swarup R, Swarup K, Penáková P, Novák O, Staswick P, Hedden P, Phillips AL, Vissenberg K, Bennett MJ, Ljung K (2016) Dioxygenase-encoding PubMed DOI PMC

Qin GJ, Gu HY, Zhao YD, Ma ZQ, Shi GL, Yang Y, Pichersky E, Chen HD, Liu MH, Chen ZL, Qu LJ (2005) An indole-3-acetic acid carboxyl methyltransferase regulates PubMed DOI PMC

Rárová L, Ncube B, Van Staden J, Fürst R, Strnad M, Gruz J (2019) Identification of narciclasine as an in vitro anti-inflammatory component of PubMed DOI

Rouleau M, Marsolais F, Richard M, Nicolle L, Voigt B, Adam G, Varin L (1999) Inactivation of brassinosteroid biological activity by a salicylate-inducible steroid sulfotransferase from PubMed DOI

Sanda S, Leustek T, Theisen MJ, Garavito RM, Benning C (2001) Recombinant PubMed DOI

Sardar P, Kempken F (2018) Characterization of indole-3-pyruvic acid pathway-mediated biosynthesis of auxin in PubMed DOI PMC

Shen Y, Diener AC (2013) PubMed DOI PMC

Sprunck S, Jacobsen HJ, Reinard T (1995) Indole-3-lactic acid is a weak auxin analogue but not an anti-auxin. J Plant Growth Regul 14:191–197 DOI

Supikova K, Kosinova A, Vavrusa M, Koplikova L, François A, Pospisil J, Zatloukal M, Wever R, Hartog A, Gruz J (2022) Sulfated phenolic acids in plants. Planta 255:124 PubMed DOI

Tarakhovskaya ER, Maslov YI, Shishova MF (2007) Phytohormones in algae. Russ J Plant Physiol 54:163–170 DOI

Torrens-Spence M, Gillaspy G, Zhao B, Harich K, White R, Li J (2012) Biochemical evaluation of a parsley tyrosine decarboxylase results in a novel 4-hydroxyphenylacetaldehyde synthase enzyme. Biochem Biophys Res Commun 418:211–216 PubMed DOI

Ugena L, Hylová A, Podlesáková K, Humplík JF, Dolezal K, De Diego N, Spíchal L (2018) Characterization of biostimulant mode of action using novel multi-trait high-throughput screening of PubMed DOI PMC

Walz A, Park S, Cohen JD, Momonoki YS, Slovin JP, Ludwig-Mueller J (2001) A gene encoding a protein modified by the phytohormone indoleacetic acid. Plant Biology (Rockville) 2001:82 PubMed PMC

Wani MC, Taylor HL, Wall ME, Coggon P, McPhail AT (1971) Plant antitumor agents.6. isolation and structure of taxol, a novel antileukemic and antitumor agent from taxus-brevifolia. J Am Chem Soc 93:2325–2327 PubMed DOI

Woodward AW, Bartel B (2005) Auxin: Regulation, action, and interaction. Ann Bot 95:707–735 PubMed DOI PMC

Xu JJ, Fang X, Li CY, Zhao Q, Martin C, Chen XY, Yang L (2018) Characterization of PubMed DOI PMC

Yi L, Dratter J, Wang C, Tunge JA, Desaire H (2006) Identification of sulfation sites of metabolites and prediction of the compounds’ biological effects. Anal Bioanal Chem 386:666–674 PubMed DOI PMC

Zazímalová E, Murphy AS, Yang HB, Hoyerová K, Hosek P (2010) Auxin transporters - why so many? Cold Spring Harb Perspect Biol 2:a001552 PubMed DOI PMC

Zhang S, van Duijn B (2014) Cellular auxin transport in algae. Plants-Basel 3:58–69 PubMed DOI PMC

Zhao YD (2010) Auxin biosynthesis and its role in plant development. Annu Rev Plant Biol 61(61):49–64 PubMed DOI PMC

Zhao YD (2012) Auxin biosynthesis: a simple two-step pathway converts tryptophan to indole-3-acetic acid in plants. Mol Plant 5:334–338 PubMed DOI PMC

Zubieta C, Ross JR, Koscheski P, Yang Y, Pichersky E, Noel JP (2003) Structural basis for substrate recognition in the salicylic acid carboxyl methyltransferase family. Plant Cell 15:1704–1716 PubMed DOI PMC