Camalexin is synthesized from indole-3-acetaldoxime, a key branching point between primary and secondary metabolism in Arabidopsis

Abstract

Characteristic for cruciferous plants is their production of N- and S-containing indole phytoalexins with disease resistance and cancer-preventive properties, previously proposed to be synthesized from indole independently of tryptophan. We show that camalexin, the indole phytoalexin of Arabidopsis thaliana, is synthesized from tryptophan via indole-3-acetaldoxime (IAOx) in a reaction catalyzed by CYP79B2 and CYP79B3. Cyp79B2/cyp79B3 double knockout mutant is devoid of camalexin, as it is also devoid of indole glucosinolates [Zhao, Y., Hull, A. K., Gupta, N. R., Goss, K. A., Alonso, J., Ecker, J. R., Normanly, J., Chory, J. & Celenza, J. L. (2002) Genes Dev. 16, 3100-3112], and isotope-labeled IAOx is incorporated into camalexin. These results demonstrate that only CYP79B2 and CYP79B3 contribute significantly to the IAOx pool from which camalexin and indole glucosinolates are synthesized. Furthermore, production of camalexin in the sur1 mutant devoid of glucosinolates excludes the possibility that camalexin is derived from indole glucosinolates. CYP79B2 plays an important role in camalexin biosynthesis in that the transcript level of CYP79B2, but not CYP79B3, is increased upon induction of camalexin by silver nitrate as evidenced by microarray analysis and promoter-beta-glucuronidase data. The structural similarity between cruciferous indole phytoalexins suggests that these compounds are biogenetically related and synthesized from tryptophan via IAOx by CYP79B homologues. The data show that IAOx is a key branching point between several secondary metabolic pathways as well as primary metabolism, where IAOx has been shown to play a critical role in IAA homeostasis.

Fig. 1.

Schematic view of biosynthetic pathways of IAOx-derived indole compounds in Arabidopsis.

Fig. 2.

Induction of camalexin in leaves of Arabidopsis after treatment with AgNO₃. Camalexin was measured in plant extracts from WT, cyp79B2, cyp79B3, and cyp79B2/cyp79B3 mutants 24 h after treatment with 5 mM AgNO₃. Black bars are 4-week-old plants (n = 6), and gray bars are 6-week-old plants (n = 10).

Fig. 3.

GUS expression in rosette leaves of 6-week-old CYP79B2p::GUS and CYP79B3p::GUS plants after AgNO₃ and P. syringae treatment. CYP79B2p::GUS plants are shown untreated (A), 8 h after spraying with 5 mM AgNO₃ (B), 16 h after spraying with 5 mM AgNO₃ (C), 16 h after infiltration with P. syringae (D), 24 h after infiltration with P. syringae (E), and 24 h after infiltration with 10 mM MgCl₂ (control) (F). CYP79B3p::GUS plants are shown untreated (G), 16 h after spraying with 5 mM AgNO₃ (H), 24 h after infiltration with P. syringae (I), and 24 h after infiltration with 10 mM MgCl₂ (control) (J). Weakly colored rings (D and E) are syringe marks.