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Benzoxazinoids: Biosynthesis and function of major defense compounds in maize VINZENZ HANDRICK
Department of Biochemistry, MPI-CE, Jena, Germany
N
O
O
O
R3 O
R1
R2
3
RECOGNITION
Herbivores
Maize response to herbivores
Plant
4
RESPONSE RECOGNITION
Herbivores
Plant
Benzoxazinoids
Maize response to herbivores
N
O
O
O
R3 O
R1
R2
Plant
5
RESPONSE
RESISTANCE
RECOGNITION
Herbivores
Plant
Benzoxazinoids
Maize response to herbivores
N
O
O
O
R3 O
R1
R2
Plant
6
Indole
NH
OPi
HOOH
Indole-3-glycerol phosphate
N
O O
O
OH
GlcOH3C
NH
DIMBOA-Glc
Monika Frey et al. Phytochemistry (2009) 70: 1645–1651.
Biosynthesis of benzoxazinoids
7
Indole
NH
OPi
HOOH
Indole-3-glycerol phosphate
N
O O
O
OH
GlcOH3C
NH
DIMBOA-Glc
Monika Frey et al. Phytochemistry (2009) 70: 1645–1651.
Stored in the vacuole of developing tissues
DIMBOA-Glc is the most abundant benzoxazinoid (BXD)
Function against different pests
BENZOXAZINOID GLYCOSIDES
Biosynthesis of benzoxazinoids
8
Indole
NH
OPi
HOOH
Indole-3-glycerol phosphate
N
O O
O
OH
GlcOH3C
NH
DIMBOA-Glc
Biosynthesis of benzoxazinoids
N
O O
O
O
GlcOH3C
CH3
HDMBOA-GlcInduced upon herbivore feeding
Gaetan Glauser et al. The Plant Journal (2011) 68: 901–911.
9
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
10
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
11
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
DIMBOA-Glc [µg*g FW-1]
Aph
id r
epro
duct
ion
[Nym
phs *
adul
t-1*w
eek-1
]
01234567
0 2000 4000
12
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
DIMBOA-Glc [µg*g FW-1]
Aph
id r
epro
duct
ion
[Nym
phs *
adul
t-1*w
eek-1
]
01234567
0 2000 4000
HDMBOA-Glc [µg*g FW-1]
Aphi
d re
prod
uctio
n
[Nym
phs *ad
ult-1
*wee
k-1]
01234567
0 2000 4000
13
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
?
DIMBOA-Glc [µg*g FW-1]
Aph
id r
epro
duct
ion
[Nym
phs *
adul
t-1*w
eek-1
]
01234567
0 2000 4000
HDMBOA-Glc [µg*g FW-1]
Aphi
d re
prod
uctio
n
[Nym
phs *ad
ult-1
*wee
k-1]
01234567
0 2000 4000
14
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
DIMBOA-Glc and HDMBOA-Glc contents correlate with aphid reproduction
?O-methyltransferase, OMT
Lisa N Meihls and Vinzenz Handrick et al. The Plant Cell (2013) 25: 2341–2355.
DIMBOA-Glc [µg*g FW-1]
Aph
id r
epro
duct
ion
[Nym
phs *
adul
t-1*w
eek-1
]
01234567
0 2000 4000
HDMBOA-Glc [µg*g FW-1]
Aphi
d re
prod
uctio
n
[Nym
phs *ad
ult-1
*wee
k-1]
01234567
0 2000 4000
15
N
O
O
O
H3CO OGlc
CH3
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
QTL mapping of candidate genes
?
16
N
O
O
O
H3CO OGlc
CH3
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
QTL mapping of candidate genes
?
17
N
O
O
O
H3CO OGlc
CH3
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
_ HDMBOA-Glc and aphid QTL comprises 3 O-methyltransferase genes
Bx10a, Bx10b, Bx10c
QTL mapping of candidate genes
?
18
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli
Candidate genes encode for active DIMBOA-Glc O-methyltransferases
19
(1) Heterologous expression of Bx10a, Bx10b and Bx10c in E. coli
(2) Purification of the recombinant Bx10a, Bx10b and Bx10c using the His-tag
Candidate genes encode for active DIMBOA-Glc O-methyltransferases
20
(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
Candidate genes encode for active DIMBOA-Glc O-methyltransferases
21
(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 HPLC-UV @ 270 nm
Candidate genes encode for active DIMBOA-Glc O-methyltransferases
22
(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 HPLC-UV @ 270 nm
FIG. HPLC chromatograms of OMT enzyme products
HDMBOA-Glc HDMBOA-Glc
HDMBOA-Glc
Candidate genes encode for active DIMBOA-Glc O-methyltransferases
Lisa N Meihls and Vinzenz Handrick et al. The Plant Cell (2013) 25: 2341–2355.
23
_ Bx10a and Bx10b are not critical for HDMBOA-Glc contents in maize
Expression analysis of DIMBOA-Glc O-methyltransferases
24
_ Bx10a and Bx10b are not critical for HDMBOA-Glc contents in maize
_ Bx10c is highly expressed in lines with high HDMBOA-Glc concentrations
Expression analysis of DIMBOA-Glc O-methyltransferases
25
Bx10c is inactivated by a transposon insertion
26
Bx10c-B73Doppia-like transposon ( ~ 8000 bp)5` 3`
Bx10c is inactivated by a transposon insertion
27
Bx10c-B73Doppia-like transposon ( ~ 8000 bp)5` 3`
_ A transposon insertion causes a natural knock out of Bx10c
Bx10c is inactivated by a transposon insertion
28
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
BX10c
DIMBOA-Glc induces callose formation
29
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
BX10c
DIMBOA-Glc induces callose formation
30
DIMBOA-Glc Callose formation
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
BX10c
DIMBOA-Glc induces 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
33DIMBOA-Glc [µg*g FW-1]
Aph
id r
epro
duct
ion
[Nym
phs *
adul
t-1*w
eek-1
]
01234567
0 2000 4000
Maize aphid resistance is caused by a transposon insertion in Bx10c
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
35
36
37
Benzoxazinoid pathway is loading …
38
Outlook
N
O
O
OH
H3CO OGlc
DIMBOA-Glc HDMBOA-Glc
N
O
O
O
H3CO OGlc
CH3
N
O
O
OH
H3CO OGlc
OCH3
N
O
O
O
H3CO OGlc
CH3
OCH3
DIM2BOA-Glc HDM2BOA-Glc
39
SINERGIA Grand of the
Jonathan Gershenzon Tobias G Köllner
Matthias Erb
Georg JanderLisa N Meihls
Gaetan Glauser
ALL CO-WORKERS
