The metabolic adaptations by which phloem-feeding insects counteract plant defense compounds are poorly known. Two-component plant defenses, such as glucosinolates, consist of a glucosylated protoxin that is activated by a glycoside hydrolase upon plant damage. Phloem-feeding herbivores are not generally believed to be negatively impacted by two-component defenses due to their slender piercing-sucking mouthparts, which minimize plant damage. However, here we document that glucosinolates are indeed activated during feeding by the whitefly Bemisia tabaci. This phloem feeder was also found to detoxify the majority of the glucosinolates it ingests by the stereoselective addition of glucose moieties, which prevents hydrolytic activation of these defense compounds. Glucosylation of glucosinolates in B. tabaci was accomplished via a transglucosidation mechanism, and two glycoside hydrolase family 13 (GH13) enzymes were shown to catalyze these reactions. This detoxification reaction was also found in a range of other phloem-feeding herbivores.
Subscribe to Journal
Get full journal access for 1 year
only $14.08 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
The datasets generated and/or analyzed during the current study are available from the corresponding authors upon reasonable request. Public databases used in the construction of the protein phylogenetic tree are provided in Supplementary Table 7 and are available from the following websites: HGSC (https://www.hgsc.bcm.edu/arthropods/colorado-potato-beetle-genome-project), BIPAA (https://bipaa.genouest.org/sp/acyrthosiphon_pisum/), Whitefly Genome Database (http://www.whiteflygenomics.org/cgi-bin/bta/index.cgi), BIPAA (https://bipaa.genouest.org/sp/myzus_persicae/), NCBI (https://www.ncbi.nlm.nih.gov/genome/annotation_euk/Bombyx_mori/101/), HGSC (https://www.hgsc.bcm.edu/arthropods/tobacco-hornworm-genome-project), NCBI (https://www.ncbi.nlm.nih.gov/genome/annotation_euk/Pieris_rapae/100/), NCBI (https://www.ncbi.nlm.nih.gov/assembly/GCA_000697945.4), Ensembl (http://metazoa.ensembl.org/Tetranychus_urticae/Info/Index) and dbCAN (http://bcb.unl.edu/dbCAN/). All other data supporting this work, if not already indicated, are available in the Supplementary Information.
Morant, A. V. et al. ß-glucosidases as detonators of plant chemical defense. Phytochemistry 69, 1795–1813 (2008).
Pentzold, S., Zagrobelny, M., Rook, F. & Bak, S. How insects overcome two-component plant chemical defence: plant ß-glucosidases as the main target for herbivore adaptation. Biol. Rev. 89, 531–551 (2014).
Tjallingii, W. F. & Esch, T. H. Fine structure of aphid stylet routes in plant tissues in correlation with EPG signals. Physiol. Entomol. 18, 317–328 (1993).
Walker, G. P. & Perring, T. M. Feeding and oviposition behavior of whiteflies (Homoptera, Aleyrodidae) interpreted from AC electronic feeding monitor wave forms. Ann. Entomol. Soc. Am. 87, 363–374 (1994).
Walling, L. L. Avoiding effective defenses: strategies employed by phloem-feeding insects. Plant Physiol. 146, 859–866 (2008).
Wang, X. W., Li, P. & Liu, S. S. Whitefly interactions with plants. Curr. Opin. Insect Sci. 19, 70–75 (2017).
De Barro, P. J., Liu, S. S., Boykin, L. M. & Dinsdale, A. B. Bemisia tabaci: a statement of species status. Annu. Rev. Entomol. 56, 1–19 (2011).
Oliveira, M. R. V., Henneberry, T. J. & Anderson, P. History, current status and collaborative research projects for Bemisia tabaci. Crop Prot. 20, 709–723 (2001).
Bones, A. M. & Rossiter, J. T. The myrosinase-glucosinolate system, its organisation and biochemistry. Physiol. Plant. 97, 194–208 (1996).
Jeschke, V., Gershenzon, J. & Vassao, D. G. Insect detoxification of glucosinolates and their hydrolysis products. Adv. Bot. Res. 80, 199–245 (2016).
Falk, K. L. & Gershenzon, J. The desert locust, Schistocerca gregaria, detoxifies the glucosinolates of Schouwia purpurea by desulfation. J. Chem. Ecol. 33, 1542–1555 (2007).
Ratzka, A., Vogel, H., Kliebenstein, D. J., Mitchell-Olds, T. & Kroymann, J. Disarming the mustard oil bomb. Proc. Natl Acad. Sci. USA 99, 11223–11228 (2002).
Malka, O. et al. Glucosinolate desulfation by the phloem-feeding insect Bemisia tabaci. J. Chem. Ecol. 42, 230–235 (2016).
Kim, J. H., Lee, B. W., Schroeder, F. C. & Jander, G. Identification of indole glucosinolate breakdown products with antifeedant effects on Myzus persicae (green peach aphid). Plant J. 54, 1015–1026 (2008).
Markovich, O. et al. Arabidopsis thaliana plants with different levels of aliphatic and indolyl-glucosinolates affect host selection and performance of Bemisia tabaci. J. Chem. Ecol. 39, 1361–1372 (2013).
Hayashi, H. & Chino, M. Chemical composition of phloem sap from the uppermost internode of the rice plant. Plant Cell Physiol. 31, 247–251 (1990).
Lohaus, G. et al. Solute balance of a maize (Zea mays L.) source leaf as affected by salt treatment with special emphasis on phloem retranslocation and ion leaching. J. Exp. Bot. 51, 1721–1732 (2000).
Haritatos, E., Keller, F. & Turgeon, R. Raffinose oligosaccharide concentrations measured in individual cell and tissue types in Cucumis melo L. leaves: implications for phloem loading. Planta 198, 614–622 (1996).
Rennie, E. A. & Turgeon, R. A comprehensive picture of phloem loading strategies. Proc. Natl Acad. Sci. USA 106, 14162–14167 (2009).
Cristofoletti, P. T., Ribeiro, A. F., Deraison, C., Rahbe, Y. & Terra, W. R. Midgut adaptation and digestive enzyme distribution in a phloem feeding insect, the pea aphid Acyrthosiphon pisum. J. Insect Physiol. 49, 11–24 (2003).
Douglas, A. E. Phloem-sap feeding by animals: problems and solutions. J. Exp. Bot. 57, 747–754 (2006).
Byrne, D. N. & Miller, W. B. Carbohydrate and amino acid composition of phloem sap and honeydew produced by Bemisia tabaci. J. Insect Physiol. 36, 433–439 (1990).
Monsan, P., Remaud-Simeon, M. & Andre, I. Transglucosidases as efficient tools for oligosaccharide and glucoconjugate synthesis. Curr. Opin. Microbiol. 13, 293–300 (2010).
Fisher, D. B., Wright, J. P. & Mittler, T. E. Osmoregulation by the aphid Myzus persicae: a physiological role for honeydew oligosaccharides. J. Insect Physiol. 30, 387–393 (1984).
Price, D. R. G. et al. Molecular characterisation of a candidate gut sucrase in the pea aphid, Acyrthosiphon pisum. Insect Biochem. Mol. Biol. 37, 307–317 (2007).
Ngiwsara, L. et al. Amino acids in conserved region II are crucial to substrate specificity, reaction velocity, and regioselectivity in the transglucosylation of honeybee GH-13 ɑ-glucosidases. Biosci. Biotechnol. Biochem. 76, 1967–1974 (2012).
Jing, X. et al. Evolutionary conservation of candidate osmoregulation genes in plant phloem sap-feeding insects. Insect Mol. Biol. 25, 251–258 (2016).
Wang, X. W. et al. Analysis of a native whitefly transcriptome and its sequence divergence with two invasive whitefly species. BMC Genomics 13, 529 (2012).
Hendrix, D. L. & Salvucci, M. E. Isobemisiose: an unusual trisaccharide abundant in the silverleaf whitefly, Bemisia argentifolii. J. Insect Physiol. 47, 423–432 (2001).
Riens, B., Lohaus, G., Heineke, D. & Heldt, H. W. Amino acid and sucrose content determined in the cytosolic, chloroplastic, and vacuolar compartments and in the phloem sap of spinach leaves. Plant Physiol. 97, 227–233 (1991).
Merritt, S. Z. Within-plant variation in concentrations of amino acids, sugar, and sinigrin in phloem sap of black mustard, Brassica nigra (L) Koch (Cruciferae). J. Chem. Ecol. 22, 1133–1145 (1996).
Nintemann, S. J. et al. Localization of the glucosinolate biosynthetic enzymes reveals distinct spatial patterns for the biosynthesis of indole and aliphatic glucosinolates. Physiol. Plant. 163, 138–154 (2018).
Andreasson, E., Jorgensen, L. B., Hoglund, A. S., Rask, L. & Meijer, J. Different myrosinase and idioblast distribution in Arabidopsis and Brassica napus. Plant Physiol. 127, 1750–1763 (2001).
Danner, H., Desurmont, G. A., Cristescu, S. M. & van Dam, N. M. Herbivore-induced plant volatiles accurately predict history of coexistence, diet breadth and feeding mode of herbivores. New Phytol. 220, 726–738 (2018).
Bak, S., Nielsen, H. L. & Halkier, B. A. The presence of CYP79 homologues in glucosinolate-producing plants shows evolutionary conservation of the enzymes in the conversion of amino acid to aldoxime in the biosynthesis of cyanogenic glucosides and glucosinolates. Plant Mol. Biol. 38, 725–734 (1998).
Eakteiman, G. et al. Targeting detoxification genes by phloem-mediated RNAi: a new approach for controlling phloem-feeding insect pests. Insect Biochem. Mol. 100, 10–21 (2018).
Luo, Y. A. et al. Towards an understanding of the molecular basis of effective RNAi against a global insect pest, the whitefly Bemisia tabaci. Insect Biochem. Mol. 88, 21–29 (2017).
Guershon, M. & Ayali, A. Innate phase behavior in the desert locust, Schistocerca gregaria. Insect Sci. 19, 649–656 (2012).
Beran, F. et al. Phyllotreta striolata flea beetles use host plant defense compounds to create their own glucosinolate-myrosinase system. Proc. Natl Acad. Sci. USA 111, 7349–7354 (2014).
Jeschke, V. et al. How glucosinolates affect generalist Lepidopteran larvae: growth, development and glucosinolate metabolism. Front. Plant Sci. 8, 1995 (2017).
Robert, C. A. M. et al. A specialist root herbivore exploits defensive metabolites to locate nutritious tissues. Ecol. Lett. 15, 55–64 (2012).
Finn, R. D. et al. The Pfam protein families database: towards a more sustainable future. Nucleic Acids Res. 44, D279–D285 (2016).
Edgar, R. C. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 32, 1792–1797 (2004).
Capella-Gutierrez, S., Silla-Martinez, J. M. & Gabaldon, T. trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25, 1972–1973 (2009).
Kalyaanamoorthy, S., Minh, B. Q., Wong, T. K. F., von Haeseler, A. & Jermiin, L. S. ModelFinder: fast model selection for accurate phylogenetic estimates. Nat. Methods 14, 587–589 (2017).
Minh, B. Q., Nguyen, M. A. T. & von Haeseler, A. Ultrafast approximation for phylogenetic bootstrap. Mol. Biol. Evol. 30, 1188–1195 (2013).
Guindon, S. et al. New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst. Biol. 59, 307–321 (2010).
Nguyen, L. T., Schmidt, H. A., von Haeseler, A. & Minh, B. Q. IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol. Biol. Evol. 32, 268–274 (2015).
Wang, X. W. et al. Transcriptome analysis and comparison reveal divergence between two invasive whitefly cryptic species. BMC Genomics 12, 458 (2011).
Bolger, A. M., Lohse, M. & Usadel, B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30, 2114–2120 (2014).
Li, B. & Dewey, C. N. RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinformatics 12, 323 (2011).
Ye, X. D. Transcriptomic analyses reveal the adaptive features and biological differences of guts from two invasive whitefly species. BMC Genomics 15, 370 (2014).
Zhang, H. et al. dbCAN2: a meta server for automated carbohydrate-active enzyme annotation. Nucleic Acids Res. 46, W95–W101 (2018).
Yin, Y. B. et al. dbCAN: a web resource for automated carbohydrate-active enzyme annotation. Nucleic Acids Res. 40, W445–W451 (2012).
Cantarel, B. L. et al. The carbohydrate-active EnZymes database (CAZy): an expert resource for Glycogenomics. Nucleic Acids Res. 37, D233–D238 (2009).
Sablok, G., Kumar, S., Ueno, S., Kuo, J. & Varotto, C. (eds) Advances in the Understanding of Biological Sciences Using Next Generation Sequencing (NGS) Approaches (Springer, 2015).
Franceus, J. & Desmet, T. Sucrose phosphorylase and related enzymes in glycoside hydrolase family 13: discovery, application and engineering. Int. J. Mol. Sci. 21, 2526 (2020).
Majzlova, K., Pukajova, Z. & Janecek, S. Tracing the evolution of the α-amylase subfamily GH13_36 covering the amylolytic enzymes intermediate between oligo-1,6-glucosidases and neopullulanases. Carbohydr. Res. 367, 48–57 (2013).
Oslancova, A. & Janecek, S. Oligo-1,6-glucosidase and neopullulanase enzyme subfamilies from the α-amylase family defined by the fifth conserved sequence region. Cell. Mol. Life Sci. 59, 1945–1959 (2002).
Stam, M. R., Danchin, E. G. J., Rancurel, C., Coutinho, P. M. & Henrissat, B. Dividing the large glycoside hydrolase family 13 into subfamilies: towards improved functional annotations of α-amylase-related proteins. Protein Eng. Des. Sel. 19, 555–562 (2006).
We thank A. Douglas (Cornell University) for the SUC1 sequence, K. Falk for assistance with graphics, the MPI-CE, DSMZ and HUJI greenhouse teams for plant and insect maintenance, and other members of the African Cassava Whitefly Project (cassavawhitefly.org) for helpful discussions. This work was supported financially by the Max Planck Society, the Deutsche Forschungsgemeinschaft (DFG Collaborative Research Center 1127 ChemBioSys) and the Natural Resources Institute, University of Greenwich from a grant provided by the Bill and Melinda Gates Foundation (OPP1058938).
The authors declare no competing interests.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
LC-MS traces of allyl-GSL, 4mtb [4-methylthiobutyl]-GSL, 3msop [3-methylsulfinylpropyl]-GSL and 4moi3m [4-methoxyindolyl-3-methyl]-GSL and their glycosides detected in the combined honeydew of 50-100 adult Bemisia tabaci MEAM1 whiteflies feeding on GSL-containing plants (kale or A. thaliana Col-0). The detected parent GSL is indicated with structure and representative color for the mass spectral trace (light gray), and the dectected subsequent glycosides represented as +162 Da and +324 Da (dark gray and black respectively).
Extended Data Fig. 2 GSL glucosylation in the whitefly Bemisia tabaci is catalyzed by a transglucosidase activity.
a, Simplified reaction mechanism of a sucrase-transglucosidase showing the two competing reaction paths: After binding of sucrose to the enzyme (A), hydrolysis of the fructose residue occurs with retention of bound glucose (B). Glucose is released (C) when sucrose concentrations are low, while transglucosidation to an acceptor (C’) occurs when acceptor concentrations are sufficiently high. This product may undergo further transglucosidation (D). b, Depiction of the results from two of the five diets not shown in Fig. 3, those diets with the 13C-labeled monosaccharides glucose and fructose. None gave labeled glycosylated GSLs, unlike feeding with sucrose labeled in the glucose portion. The results are consistent with a transglucosidase activity that initially hydrolyzes sucrose and links the resulting glucose moiety to the plant GSL.
Extended Data Fig. 3 Maximum likelihood circular cladogram showing the relationship of glycoside hydrolase family 13 enzymes from nine chosen herbivore species.
The tree was inferred using a total of 205 sequences. Ultrafast bootstrap45 and Shimodaira–Hasegawa approximate likelihood ratio test (SH-aLRT)46 validation values lower than 95 are presented close to the corresponding nodes. GH13 members from Bemisia tabaci are highlighted by red nodes. SUC1–5 are indicated by bold text in red. Colors surrounding the cladogram indicate the feeding guild of the corresponding species. Thin colored inner circles specify the subfamily of the corresponding GH13 enzymes. Subfamilies with less than four proteins are marked only by the subfamily number (for more details see Supplementary Table 8). The protein sequences are named according to their GenBank accession numbers, or their names in the released proteome. Species name abbreviations are as indicated: Ld, Leptinotarsa decemlineata; ACYP, Acyrthosiphon pisum; BT, Bemisia tabaci; MPER, Myzus persicae; Bm, Bombyx mori; Msex, Manduca sexta; Prapae, Pieris rapae; FOCC, Frankliniella occidentalis; Tetur, Tetranychus urticae.
About this article
Cite this article
Malka, O., Easson, M.L.A.E., Paetz, C. et al. Glucosylation prevents plant defense activation in phloem-feeding insects. Nat Chem Biol 16, 1420–1426 (2020). https://doi.org/10.1038/s41589-020-00658-6