2. Benzoxazinoids: Biosynthesis and function
of major defense compounds in maize
VINZENZ HANDRICK
Department of Biochemistry, MPI-CE, Jena, Germany

3. Maize response to herbivores
RECOGNITION
Herbivores
Plant
BXD Biosynthesis

4. Maize response to herbivores
RECOGNITION
Herbivores
Plant
Benzoxazinoids
Plant
BXD Biosynthesis

5. Maize response to herbivores
RESISTANCE
RECOGNITION
Herbivores
Plant
Benzoxazinoids
Plant
BXD Biosynthesis

6. Indole-3-glycerol phosphate
Biosynthesis of benzoxazinoids
Indole-3-glycerol phosphate
Indole
DIMBOA-Glc
Monika Frey et al. Phytochemistry (2009) 70: 1645–1651.

7. Indole-3-glycerol phosphate
Biosynthesis of benzoxazinoids
Indole-3-glycerol phosphate
Indole
DIMBOA-Glc
BENZOXAZINOID GLYCOSIDES
Stored in the vacuole of developing tissues
DIMBOA-Glc is the most abundant benzoxazinoid (BXD)
Function against different pests
Monika Frey et al. Phytochemistry (2009) 70: 1645–1651.

8. Indole-3-glycerol phosphate
Biosynthesis of benzoxazinoids
Indole-3-glycerol phosphate
Indole
DIMBOA-Glc
Induced upon herbivore feeding
HDMBOA-Glc
Gaetan Glauser et al. The Plant Journal (2011) 68: 901–911.

9. DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
BXD Biosynthesis

10. DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
BXD Biosynthesis

11. DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
DIMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
BXD Biosynthesis

12. DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
DIMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
HDMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
BXD Biosynthesis

13. ? DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
DIMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
HDMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
BXD Biosynthesis

14. ? DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
O-methyltransferase, OMT
DIMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
HDMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000
Lisa N Meihls and Vinzenz Handrick et al. The Plant Cell (2013) 25: 2341–2355.
BXD Biosynthesis

15. QTL mapping of candidate genes
DIMBOA-Glc
HDMBOA-Glc ?

16. QTL mapping of candidate genes
DIMBOA-Glc
HDMBOA-Glc ?

17. QTL mapping of candidate genes
DIMBOA-Glc
HDMBOA-Glc ?
HDMBOA-Glc and aphid QTL comprises 3 O-methyltransferase genes
Bx10a, Bx10b, Bx10c

18. Candidate genes encode for active DIMBOA-Glc O-methyltransferases
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli

19. Candidate genes encode for active DIMBOA-Glc O-methyltransferases
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli
(2) Purification of the recombinant Bx10a, Bx10b and Bx10c using the His-tag

20. Candidate genes encode for active DIMBOA-Glc O-methyltransferases
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli
(2) Purification of the recombinant Bx10a, Bx10b and Bx10c using the His-tag
(3) In vitro enzyme assays with DIMBOA-Glc and the methyl group donor SAM

21. Candidate genes encode for active DIMBOA-Glc O-methyltransferases
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli
(2) Purification of the recombinant Bx10a, Bx10b and Bx10c using the His-tag
(3) In vitro enzyme assays with DIMBOA-Glc and the methyl group donor SAM
(4) Detection of BX10 products by 270 nm

22. Candidate genes encode for active DIMBOA-Glc O-methyltransferases
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli
(2) Purification of the recombinant Bx10a, Bx10b and Bx10c using the His-tag
(3) In vitro enzyme assays with DIMBOA-Glc and the methyl group donor SAM
(4) Detection of BX10 products by 270 nm
FIG. HPLC chromatograms of OMT enzyme products
HDMBOA-Glc
Lisa N Meihls and Vinzenz Handrick et al. The Plant Cell (2013) 25: 2341–2355.

23. Expression analysis of DIMBOA-Glc O-methyltransferases
Bx10a and Bx10b are not critical for HDMBOA-Glc contents in maize

24. Expression analysis of DIMBOA-Glc O-methyltransferases
Bx10a and Bx10b are not critical for HDMBOA-Glc contents in maize
Bx10c is highly expressed in lines with high HDMBOA-Glc concentrations

25. Bx10c is inactivated by a transposon insertion

26. Bx10c is inactivated by a transposon insertion
Bx10c-B73
Doppia-like transposon ( ~ 8000 bp) 5` 3`

27. Bx10c is inactivated by a transposon insertion
Bx10c-B73
Doppia-like transposon ( ~ 8000 bp) 5` 3`
A transposon insertion causes a natural knock out of Bx10c

28. DIMBOA-Glc induces callose formation
HDMBOA-Glc
BX10c

29. DIMBOA-Glc induces callose formation
HDMBOA-Glc
BX10c

30. DIMBOA-Glc induces callose formation
HDMBOA-Glc
BX10c
DIMBOA-Glc
Callose formation

31. Maize aphid resistance is caused by a transposon insertion in Bx10c

32. Maize aphid resistance is caused by a transposon insertion in Bx10c

33. Maize aphid resistance is caused by a transposon insertion in Bx10c
DIMBOA-Glc [µg*g FW-1]
[Nymphs*adult-1*week-1]
Aphid reproduction 1 2 3 4 5 6 7
2000
4000

34. Maize aphid resistance is caused by a transposon insertion in Bx10c
_ The Bx10c transposon insertion and the resulting DIMBOA-Glc accumulation lead to an increase in callose deposition and leaf aphid resistance

37. Benzoxazinoid pathway is loading …

38. Outlook
DIMBOA-Glc
HDMBOA-Glc
DIM2BOA-Glc
HDM2BOA-Glc

39.  ALL CO-WORKERS SINERGIA Grand of the Jonathan Gershenzon
Tobias G Köllner
Lisa N Meihls
Georg Jander
SINERGIA Grand of the
Gaetan Glauser
Matthias Erb
ALL CO-WORKERS
BXD Biosynthesis