Article
Reference
Spatially and genetically distinct control of seed germination by phytochromes A and B
LEE, Keun Pyo, et al.
Abstract
Phytochromes phyB and phyA mediate a remarkable developmental switch whereby, early upon seed imbibition, canopy light prevents phyB-dependent germination, whereas later on, it stimulates phyA-dependent germination. Using a seed coat bedding assay where the growth of dissected embryos is monitored under the influence of dissected endosperm, allowing combinatorial use of mutant embryos and endosperm, we show that canopy light specifically inactivates phyB activity in the endosperm to override phyA-dependent signaling in the embryo. This interference involves abscisic acid (ABA) release from the endosperm and distinct spatial activities of phytochrome signaling components. Under the canopy, endospermic ABA opposes phyA signaling through the transcription factor (TF) ABI5, which shares with the TF PIF1 several target genes that negatively regulate germination in the embryo. ABI5 enhances the expression of phytochrome signaling genes PIF1, SOMNUS, GAI, and RGA, but also of ABA and gibberellic acid (GA) metabolic genes. Over time, weaker ABA-dependent responses eventually enable phyA-dependent germination, a distinct type of [...]
LEE, Keun Pyo, et al . Spatially and genetically distinct control of seed germination by phytochromes A and B. Genes and Development , 2012, vol. 26, no. 17, p. 1984-1996
DOI : 10.1101/gad.194266.112 PMID : 22948663
Available at:
http://archive-ouverte.unige.ch/unige:43509
Disclaimer: layout of this document may differ from the published version.
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Lee_Sup.Fig1.
PHYA PHYB Loading
S S E C WT phyA/B FR pulse
0 4 Time (h)
Seed (S) Dissection at 4h
Coat (C) Embryo (E)
A B
WT (ft-1) seeds
Coat-less embryos
BCDE
ACDE
ACDE (+GA)
1
2
Length (mm) 3 0 1 2 3 4
1 2 3 4
1 mm
FR pulse
0 4 50
Time (h)
*Diss. at 4h
50h 4h
C
Relative transcript (/UBQ11)
0 1.0 2.0
0 0.7 1.4
GA3ox1 GA3ox2
0 120 240
0 140 280
NCED6 NCDE9
FR pulse
0 4 28
(h)
FR pulse
0 4 28
(h)
Seed Coat
Embryo
Embryo Coat
Relative transcript (/UBQ11)
D
Relative transcript (/UBQ11)
0.000 0.004
ABi4 EPR1
0.300 0.600 0.900 1.200
Time (h)
0 4 48
(h) FR
12 24
Dissection at each time points
4 12 24 48 48
WT phyA
PHYA Loading COAT
B
Lee_Sup.Fig2.
0 5 10 15 20 25
0 10 20 30 40 FR FR/FR
COAT EMBRYO
ABA levels (pmol/g.f.w.) FR FR/FR
0 4 88
(h) 48
FR FR/FR
Dissection
C
ABA levels (pmol/g.f.w.)
Length (mm)
0 1 2 3
1 2 3 4
A
Lee_Sup.Fig3.
- ABA
+ ABA
0 1 2 3 4
1 2 1
2
0 0.4 0.8 1.2
1 2 0 0.4 0.8 1.2
1 2 FR
0 4 50
(h)
Length (mm)
GA3ox1 GA3ox2
Rel. transcript (/UBQ11) 1mm
0 40 80
2 3 0 40 80
2 3 GA3ox1 GA3ox2
Relative transcript in EMBRYO(/UBQ11)
B A
Rel. transcript (/UBQ11)
C
Germination (%)
0 4 120
(h) 12 FR FR/FR
0 20 40 60 80 100
WT abi5 phyA
D
0 5
10 GA3ox1
0 25
50 GA3ox2 FR/FR
0 4 36
(h) 12 FR
Rel. transcript (/UBQ11)
Lee_Sup.Fig4.
0 0.6 1.2
0 48 96 144 192
EMBRYO COAT
0 0.6 1.2
0 48 96 144 192 NCED6
NCED9
0 0.6 1.2
0 48 96 144 192
0 0.6 1.2
0 48 96 144 192 ABI3
ABI5
0 4 144 192
FR
96 48 24 12
Time after imbibition (h)
0 0.6 1.2
0 48 96 144 192
0 0.6 1.2
0 48 96 144 192 GA3ox1
GA3ox2
B
0 0.6 1.2
0 48 96 144 192 GA3ox2 0
0.6 1.2
0 48 96 144 192 GA3ox1
0 4 192
Time (h) 144
FR
96 48 24 12
SEED
A
Time after imbibition (h)
Rel. transcript (/UBQ11)
Relative transcript (/PP2A)
0 0.6 1.2
0 48 96 144 192 NCED9 0
0.6 1.2
0 48 96 144 192 NCED6
0 0.6 1.2
0 48 96 144 192
0 0.6 1.2
0 48 96 144 192 ABI3
ABI5
Relative transcript (/PP2A)
0 0.6 1.2
0 48 96 144 192
0 0.6 1.2
0 48 96 144 192 GA3ox1
GA3ox2
FR/R, 28h
FR/R, 28h
FR/R, 28h
FR/R, 28h
FR/R, 28h
FR/R, 28h FR/R,
28h
FR/R, 28h FR/R,
28h
FR/R, 28h
WT
pif1
som
0.5 mm
Relative transcript (/UBQ11)
0 10 20 0 2.0 4.0
0 1.5 3.0
0 2.5 5.0 GA3ox1
GA3ox2 EMBRYO COAT
0 0.7 1.4
0 0.6 1.2
0 0.6 1.2
0 1.2 2.4 NCED6
NCED9
*Diss.
0 1 2
0 2 4 NCED9
0 5 10 GA3ox1
0 6 12 GA3ox2 Relative transcript in COAT(/UBQ11)
NCED6
U*
FR
0 4 28
D
(h)0 0.5 1.0
Relative transcript in COAT(/UBQ11)
WT pif1 SOM
0 10 20
0 60 120
GA3ox1
GA3ox2
Relative transcript in EMBRYO(/UBQ11)
0 0.6 1.2
0 0.6 1.2
NCED6
NCED9 U*
*Diss.
0 4 72
(h) FR
0 4 32
(h) FR
*Diss.
0 4 72
(h) FR
*Diss.
0 4 72
(h) FR
A B C E
Lee_Sup.Fig5.
C
Gene Peak id Width Fold change p-value G-box [G/T][A/C]CACGTG[G/T][C/A]
GA3ox1 I 250 9.1 0.018 no no
GA3ox2 I 1150 7.3 0.011 yes yes
II 250 6.3 0.031 no no
NCED6 I 150 4.3 0.058 no no
NCED9 I 725 11.5 0.005 yes no
II 325 4.2 0.051 no no
PIF1 I 750 3.6 0.051 no no
SOM I 350 10.5 0.011 yes no
II 325 12.1 0.009 no no
GAI
I 250 11.5 0.011 no no
II 525 13.2 0.005 yes yes
III 300 9.0 0.014 no no
RGA I 450 8.7 0.011 no yes
RGL2 I 350 10.2 0.010 yes no
II 400 7.6 0.018 no no
B
GA3ox1 GA3ox2
NCED6 NCED9
PIF1 SOM
GAI RGA
RGL2
Relative transcript(/UBQ11)
0 4 28
Time (h) FR
0 1.5 3.0
0 1.5 3.0
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
A
0 4.0 8.0
0 6 12
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 3.5 7.0
0 7 14
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2
0 0.6 1.2 GA3ox1 GA3ox2
NCED6 NCED9
PIF1 SOM
GAI RGA
RGL2
Relative transcript (/UBQ11)
GA3ox1 GA3ox2
NCED6 NCED9
PIF1 SOM
GAI RGA
RGL2
Relative transcript (/UBQ11)
0 4 72
Time (h) FR
Seed Coat
Embryo
EMBRYO COAT SEED
Lee_Sup.Fig6.
Supplemental Table 1. List of primer sequences used for qRT-PCR in this study
Gene Accession Nr. Forward Primer Reverse Primer
GA3ox1 At1g15550 5'-CGGTGAAAACATCCCTCTCATC-3' 5'-TACATGCATGACCGATTTGGTT-3' GA3ox2 At1g80340 5'-ACTCGACCCTCATGACAATTCTG-3' 5'-CCACGTCATCCCGGAAAAC-3' NCED6 At3g24220 5'-TTGAGTCCTCTATCGTTATTCCTATGG-3' 5'-GGAGCGAAGTTACCTGATAATTGAA-3' NCED9 At1g78390 5'-GGAAAACGCCATGATCTCACA-3' 5'-GCAGGATCCGCCGTTTTAG-3' ABI3 At3g24650 5'-AACCAATACGCATCAGGAAACTG-3' 5'-AAACCTTGACTGCTGTCTAATGGA-3' ABI4 At2g40220 5’-TGGGTCTGGATCCGGGTTATA-3’ 5’-CCTCCAAAAGGCCAAATGG-3’
ABI5 At2g36270 5'-AAGAGGAAGCAACAGTATTTTGAGAGT-3' 5'-CGCAATCTCCCGTTCGATT-3' PIF1 At2g20180 5'-TGTCAATGGGATGTGGAATGA-3' 5'-CATCGCCATATGAGGCATGTA-3' SOM At1g03790 5'-TCCGGATGTTCGAATTCAAGAT-3' 5'-GCAAAAGGACAATCAGTCCAATC-3' GAI At1g14920 5'-CCCGGTGACGCGATTCT-3' 5'-TCCGCCGCCTTGGTTAG-3'
RGA At2g01570 5'-CTCTGAGCTTAATCCTCCTCCTCTT-3' 5'-CATAATCCGAAGCCGGAAAAC-3' RGL2 At3g03450 5'-GTTGTGTGGATAGATCCGAATACG-3' 5'-TTCCTCGTCAAAGACTTCCTCTTC-3' EPR1 At2g27380 5’-TCTCTTAGTGGTGCTTCAGTTGCT-3’ 5’-GCGTGACCACTATCAGTCTCATACTT-3’
UBQ11 At4g05050 5'-TTCATTTGGTCTTGCGTCTG-3' 5'-GAAGATGAGACGCTGCTGGT-3' PP2A At1g13320 5'-GGACCGGAGCCAACTAGGA-3' 5'-GCTATCCGAACTTCTGCCTCATT-3'
