The role of two GLYCOALKALOID METABOLISM genes in α-tomatine biosynthesis and basal defense in tomato

You, Y.; Balaji, A.; Herrera Valderrama, A. L.; Denarie, M.-E.; Hassan Muralidhar, S.; Ramirez Gaona, M.; Hanika, K.; Verstappen, F.; Kappers, I. F.; van Kan, J. A. L.

doi:10.64898/2026.02.16.705890

bioRxiv · 10.64898/2026.02.16.705890

The role of two GLYCOALKALOID METABOLISM genes in α-tomatine biosynthesis and basal defense in tomato

You, Y.·Balaji, A.·Herrera Valderrama, A. L.·Denarie, M.-E.·Hassan Muralidhar, S.·Ramirez Gaona, M.·Hanika, K.·Verstappen, F.·Kappers, I. F.·van Kan, J. A. L.

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Abstract

Steroidal glycoalkaloids and saponins are plant cholesterol-based steroid metabolites with antimicrobial activities and potential pharmacological value. The saponin uttroside B from black nightshade (Solanum nigrum) plays an important role in defense against herbivorous insect and exhibits anti-hepatocellular carcinoma activity. The tomato (S. lycopersicum) glycoalkaloid -tomatine has been studied because of its antinutritional effects, however, its role in protecting plants from fungal pathogens remains understudied. The biosynthetic pathway of -tomatine involves multiple clustered genes designated as glycoalkaloid metabolism (GAME) genes. In this study, we generated single knockout mutants of SlGAME4 and SlGAME2 by CRISPR/Cas9-based genome editing. The SlGAME4 mutants did not accumulate glycoalkaloids but instead redirected resources towards steroidal saponin (uttroside B) synthesis. SlGAME2 mutants contained unaltered -tomatine contents indicating that the SlGAME2 gene, previously reported to catalyze the transfer of xylose to {beta}1-tomatine, is not involved in -tomatine biosynthesis. Infection assays with four fungal tomato pathogens demonstrated that the SlGAME4 mutant plants were slightly more susceptible to Botrytis cinerea, but equally susceptible to the other three fungi. Up-regulation of -tomatine-responsive genes in B. cinerea was observed during infection on SlGAME4 mutant tomato, as well as on S. nigrum suggesting that uttroside B induces a fungal transcriptional response similar to -tomatine. Furthermore, we observed that tolerance mechanisms to plant saponins mediated by glycosyl hydrolases and a glycosyltransferase contribute to virulence of B. cinerea on SlGAME4 mutant plants and S. nigrum. This indicates that also uttroside B contributes to defense against fungal pathogens and can be detoxified by B. cinerea.

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You, Y., Balaji, A., Herrera Valderrama, A. L., Denarie, M.-E., Hassan Muralidhar, S., Ramirez Gaona, M., Hanika, K., Verstappen, F., Kappers, I. F., van Kan, J. A. L.. 2026-02-17. The role of two GLYCOALKALOID METABOLISM genes in α-tomatine biosynthesis and basal defense in tomato. https://doi.org/10.64898/2026.02.16.705890

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