title: UDP-glycosyltransferases from the UGT73C subfamily in Barbarea vulgaris catalyze sapogenin 3-O-glucosylation in saponin-mediated insect resistance.
abstract: Triterpenoid saponins are bioactive metabolites that have evolved recurrently in plants, presumably for defense. Their biosynthesis is poorly understood, as is the relationship between bioactivity and structure. Barbarea vulgaris is the only crucifer known to produce saponins. Hederagenin and oleanolic acid cellobioside make some B. vulgaris plants resistant to important insect pests, while other, susceptible plants produce different saponins. Resistance could be caused by glucosylation of the sapogenins. We identified four family 1 glycosyltransferases (UGTs) that catalyze 3-O-glucosylation of the sapogenins oleanolic acid and hederagenin. Among these, UGT73C10 and UGT73C11 show highest activity, substrate specificity and regiospecificity, and are under positive selection, while UGT73C12 and UGT73C13 show lower substrate specificity and regiospecificity and are under purifying selection. The expression of UGT73C10 and UGT73C11 in different B. vulgaris organs correlates with saponin abundance. Monoglucosylated hederagenin and oleanolic acid were produced in vitro and tested for effects on P. nemorum. 3-O-β-d-Glc hederagenin strongly deterred feeding, while 3-O-β-d-Glc oleanolic acid only had a minor effect, showing that hydroxylation of C23 is important for resistance to this herbivore. The closest homolog in Arabidopsis thaliana, UGT73C5, only showed weak activity toward sapogenins. This indicates that UGT73C10 and UGT73C11 have neofunctionalized to specifically glucosylate sapogenins at the C3 position and demonstrates that C3 monoglucosylation activates resistance. As the UGTs from both the resistant and susceptible types of B. vulgaris glucosylate sapogenins and are not located in the known quantitative trait loci for resistance, the difference between the susceptible and resistant plant types is determined at an earlier stage in saponin biosynthesis.
authors.affiliation: Department of Plant Biology and Biotechnology, University of Copenhagen, 1871 Frederiksberg, Denmark.
authors.forename: Jörg M
authors.initials: JM
authors.lastname: Augustin
authors.forename: Sylvia
authors.initials: S
authors.lastname: Drok
authors.forename: Tetsuro
authors.initials: T
authors.lastname: Shinoda
authors.forename: Kazutsuka
authors.initials: K
authors.lastname: Sanmiya
authors.forename: Jens Kvist
authors.initials: JK
authors.lastname: Nielsen
authors.forename: Bekzod
authors.initials: B
authors.lastname: Khakimov
authors.forename: Carl Erik
authors.initials: CE
authors.lastname: Olsen
authors.forename: Esben Halkjær
authors.initials: EH
authors.lastname: Hansen
authors.forename: Vera
authors.initials: V
authors.lastname: Kuzina
authors.forename: Claus Thorn
authors.initials: CT
authors.lastname: Ekstrøm
authors.forename: Thure
authors.initials: T
authors.lastname: Hauser
authors.forename: Søren
authors.initials: S
authors.lastname: Bak
mesh_terms: D000818:Animals; D031216:Barbarea / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D055162:Biocatalysis; D015870:Gene Expression; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D060434:Herbivory; D007313:Insecta / Q000502:physiology*; D007700:Kinetics; D009828:Oleanolic Acid / Q000031:analogs & derivatives; D009928:Organ Specificity / Q000235:genetics; D010802:Phylogeny; D018515:Plant Leaves / Q000378:metabolism; D012502:Sapogenins / Q000378:metabolism*; D012503:Saponins / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D014530:Uridine Diphosphate / Q000378:metabolism*
chemical_list: C000609702:3-O-oleanolic acid monoglucoside; D012502:Sapogenins; D012503:Saponins; D014530:Uridine Diphosphate; D009828:Oleanolic Acid; D005964:Glucosyltransferases; C025763:hederagenin