f Transcription factors in abiotic stress response Transcription ... - ITQB
f Transcription factors in abiotic stress response Transcription ... - ITQB
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<strong>Transcription</strong> <strong>factors</strong><br />
<strong>in</strong> <strong>abiotic</strong> <strong>stress</strong> <strong>response</strong><br />
Nelson Saibo<br />
<strong>ITQB</strong><br />
ADONIS, 6 th March 2009<br />
Outl<strong>in</strong>e<br />
1 - Plant <strong>response</strong>s to adverse environmental conditions<br />
2 - <strong>Transcription</strong> <strong>factors</strong> and transcriptional regulation<br />
3 - <strong>Transcription</strong> <strong>factors</strong> <strong>in</strong>volved <strong>in</strong> <strong>abiotic</strong> <strong>stress</strong> <strong>response</strong>s<br />
4 -TFs and phtosynthetic <strong>response</strong>s to <strong>abiotic</strong> <strong>stress</strong><br />
5 - Identification and characterization of novel TFs<br />
ADONIS, 6 th March 2009<br />
1
Plants and the environmental conditions<br />
Environment altered beyond its normal<br />
range of variation to adversely affect<br />
the <strong>in</strong>dividual physiology of the<br />
organism <strong>in</strong> a significant way<br />
Plant <strong>stress</strong><br />
ADONIS, 6 th March 2009<br />
Plant <strong>response</strong> to <strong>abiotic</strong> <strong>stress</strong><br />
Cold Drought<br />
Sal<strong>in</strong>ity<br />
Chemical<br />
pollution<br />
Heat<br />
Cell damage<br />
Secondary <strong>stress</strong>:<br />
Osmotic <strong>stress</strong><br />
Oxidative <strong>stress</strong><br />
Signal sens<strong>in</strong>g, perception and transduction<br />
ABIOTIC STRESS<br />
50% crop loss world wide<br />
Osmosensor, second mensagers,<br />
MAPKs, Ca 2+ sensors, CDPKs<br />
<strong>Transcription</strong> control<br />
TFs: CBF/DREB, bZIB, MYC/MYB…<br />
Gene activation<br />
Osmoprotection, water and ion movement,<br />
detoxification, and chaperone functions<br />
Recovery of cellular homeostasis,<br />
functional and structural protection of<br />
prote<strong>in</strong>s and membranes<br />
Stress tolerance<br />
ADONIS, 6 th March 2009<br />
2
Abiotic <strong>stress</strong> tolerance is a<br />
multigenic trait<br />
Environmental stimuli<br />
or <strong>stress</strong><br />
TF<br />
Stress responsive genes<br />
STRESS TOLERANCE<br />
ADONIS, 6 th March 2009<br />
<strong>Transcription</strong> Factors<br />
<strong>Transcription</strong> <strong>factors</strong> (TFs) - prote<strong>in</strong>s that show sequence-specific<br />
DNA-b<strong>in</strong>d<strong>in</strong>g and that are capable acivat<strong>in</strong>g or repress<strong>in</strong>g gene transcription.<br />
+1<br />
Basic<br />
<strong>Transcription</strong><br />
mach<strong>in</strong>ery<br />
GGCATGGC TATA gene<br />
mRNA<br />
Promoter<br />
<strong>Transcription</strong> coregulators (coactivators/corepressors), chromat<strong>in</strong><br />
remodelers, histone acetylases, k<strong>in</strong>ases, and methylases play crucial<br />
roles <strong>in</strong> gene regulation, but lack DNA bid<strong>in</strong>g doma<strong>in</strong>s and therefore are<br />
not classified as TFs .<br />
ADONIS, 6 th March 2009<br />
3
<strong>Transcription</strong> <strong>in</strong> Eukaryotes<br />
a<br />
d<br />
b<br />
e<br />
c<br />
f<br />
ADONIS, 6 th March 2009<br />
<strong>Transcription</strong> <strong>in</strong> Eukaryotes<br />
Basic transcriptional<br />
mach<strong>in</strong>ery<br />
ADONIS, 6 th March 2009<br />
4
Schematic diagram of a prototypical<br />
transcription factor<br />
TFs conta<strong>in</strong> DNA-b<strong>in</strong>d<strong>in</strong>g doma<strong>in</strong> (DBD), signal sens<strong>in</strong>g doma<strong>in</strong> (SSD),<br />
and a transactivation doma<strong>in</strong> (TAD)<br />
The order of placement and the number of doma<strong>in</strong>s may differ <strong>in</strong> various<br />
types of TFs<br />
The transactivation and signal sens<strong>in</strong>g functions are frequently conta<strong>in</strong>ed<br />
with<strong>in</strong> the same doma<strong>in</strong><br />
ADONIS, 6 th March 2009<br />
<strong>Transcription</strong> Factor Families<br />
DNA b<strong>in</strong>d<strong>in</strong>g doma<strong>in</strong><br />
FAMILY<br />
DNA b<strong>in</strong>d<strong>in</strong>g doma<strong>in</strong>(s)<br />
+<br />
(prote<strong>in</strong>-prote<strong>in</strong><br />
<strong>in</strong>teraction doma<strong>in</strong>s)<br />
Subfamily<br />
ADONIS, 6 th March 2009<br />
5
Arabidopsis<br />
<strong>Transcription</strong> Factors<br />
Relationships and doma<strong>in</strong><br />
shuffl<strong>in</strong>g among the different<br />
Arabidopsis transcription<br />
factor families<br />
Riechmann (2000) Science 290, 2105<br />
ADONIS, 6 th March 2009<br />
Eukaryotic <strong>Transcription</strong>al Regulators<br />
ADONIS, 6 th March 2009<br />
6
Plant TF families and their function<br />
AP2/ERF (144) Development (flower/seed/root); metabolic pathways;<br />
<strong>stress</strong> <strong>response</strong>; hormone <strong>response</strong> (ABA/C 2 H 4 )<br />
bHLH (139) Development (trichome/root/carpel) <strong>abiotic</strong> <strong>stress</strong>;<br />
secondary metabolism; light <strong>response</strong>s;<br />
MYB (190) Development; secondary metabolism; defence <strong>response</strong>;<br />
<strong>abiotic</strong> <strong>stress</strong>; hormone <strong>response</strong> (ABA/GA 3 ); cell cycle; light<br />
C2H2(Zn) (112) Flower/seed development; <strong>abiotic</strong> <strong>stress</strong>; light<br />
NAC (109) Development (meristem); aux<strong>in</strong>-<strong>response</strong>; virus resistance; *<br />
HB (90) Development (several); sucrose signall<strong>in</strong>g; cell death; *<br />
MADS (82) Reproductive organs development; flower<strong>in</strong>g time/abscission; *<br />
bZIP (77) Flower/leaf/photomorphogenic development; seed-storage;<br />
WRKY (72)<br />
defence <strong>response</strong>; hormone <strong>response</strong>/biosynthesis; *<br />
Defence <strong>response</strong>; *<br />
C2C2(Zn) (104) Seed development/metabolism; flower<strong>in</strong>g time; circadian<br />
rhythm; *<br />
* Abiotic <strong>stress</strong><br />
Riechmann 2002 <strong>Transcription</strong>al regulation: an overview. The Arabidopsis Book<br />
ADONIS, 6 th March 2009<br />
<strong>Transcription</strong>al regulatory network<br />
Environmental stimuli<br />
or <strong>stress</strong><br />
Developmental signals<br />
Induction<br />
P<br />
Modification<br />
U<br />
S<br />
A<br />
TF1<br />
Activ.<br />
B<br />
TF2 P<br />
C<br />
TF3<br />
U S<br />
promoter<br />
TFIID<br />
TFIIA<br />
TF TF TF<br />
II IIF II<br />
H<br />
P<br />
E<br />
TFIIB RNA pol II<br />
Basic mach<strong>in</strong>ery<br />
<strong>Transcription</strong><br />
TATA +1 ATGNNNNN<br />
gene<br />
mRNA<br />
ADONIS, 6 th March 2009<br />
7
Abiotic <strong>stress</strong> transcriptional network<br />
Biotic <strong>stress</strong><br />
and wound<strong>in</strong>g<br />
Drought, High sal<strong>in</strong>ity<br />
Signal perception<br />
Cold<br />
Jasmonic<br />
acid<br />
ABA<br />
ABA-<strong>in</strong>dependent<br />
ICE1<br />
HOS1<br />
SIZ1<br />
MYB15<br />
ICE1<br />
MYB<br />
MYC<br />
AREB/ABF<br />
ZF-HD<br />
NAC<br />
DREB2<br />
?<br />
CBF4/DREB1D<br />
CBF3/DREB1A<br />
CBF1/DREB1B<br />
?<br />
DREB2<br />
ZAT12<br />
AREB/ABF<br />
CBF2/DREB1C<br />
MYCR<br />
RD22<br />
MYBR<br />
ABRE<br />
rps-1like NACR<br />
RD29B<br />
ERD1<br />
CAB<br />
?<br />
STZ/ZAT10<br />
DRE/CRT<br />
RD29A<br />
? ?<br />
Annals Botany 2009 103, 609<br />
ADONIS, 6 th March 2009<br />
Manipulation of TFs to improve<br />
<strong>abiotic</strong> <strong>stress</strong> tolerance<br />
Nature Biotechnology 1999 17, 287<br />
ADONIS, 6 th March 2009<br />
8
DREB1A driven by the 35 S CaMV vs<br />
<strong>stress</strong> <strong>in</strong>ducible rd29A promoter<br />
35S:TF growth retardation<br />
Nature Biotechnology 1999 17, 287<br />
ADONIS, 6 th March 2009<br />
Both 35S:DREB1A and rd29A:DREB1A<br />
show enhanced <strong>stress</strong> tolerance<br />
Nature Biotechnology 1999 17, 287<br />
ADONIS, 6 th March 2009<br />
9
DREB1A target genes are strongly<br />
expressed under control conditions<br />
OX mimics i acclimation<br />
Higher <strong>stress</strong> tolerance<br />
Nature Biotechnology 1999 17, 287<br />
ADONIS, 6 th March 2009<br />
Overexpression of HvCBF4 <strong>in</strong> rice enhances<br />
drought tolerance<br />
NT<br />
Ubq::<br />
::HvCBF4<br />
12 days<br />
drought<br />
1 week<br />
recover<strong>in</strong>g<br />
Survival rate:<br />
• Plants overexpress<strong>in</strong>g HvCBF4 - 90%<br />
• Non transformed plants– 19%<br />
ADONIS, 6 th March 2009<br />
10
Overexpression of HvCBF4 <strong>in</strong> rice enhances<br />
drought tolerance<br />
Ubi::HvCBF4<br />
AtRD29A::HvCBF4<br />
NT<br />
0 2 3 4 6 8 9<br />
Days after withhold water<br />
0 2 4 6 8 11<br />
Days after withhold water<br />
ADONIS, 6 th March 2009<br />
AtDREB1A overexpressed <strong>in</strong> chrysanthemum<br />
enhances tolerance to heat <strong>stress</strong><br />
Survival rate:<br />
70,8%<br />
16,3%<br />
36h at 45ºC<br />
3 weeks 22ºC<br />
Plant Mol Biol<br />
DOI 10.1007/s11103-009-9468-z<br />
ADONIS, 6 th March 2009<br />
11
Higher photosynthetic capacity and<br />
elevated activity of Rubisco<br />
Plant Mol Biol<br />
DOI 10.1007/s11103-009-9468-z<br />
ADONIS, 6 th March 2009<br />
TF overexpression can improve<br />
photosynthetic performance under<br />
<strong>abiotic</strong> <strong>stress</strong><br />
ADONIS, 6 th March 2009<br />
12
Photosynthetic <strong>response</strong>s<br />
to <strong>abiotic</strong> <strong>stress</strong><br />
ABIOITIC STRESS<br />
Stomatal control of CO 2 diffusion<br />
Photosystem II repair<br />
Electron transport<br />
Rubisco activity<br />
Scaveng<strong>in</strong>g of ROS<br />
Photorespiration<br />
Photosynthetic efficiency<br />
is greatly decreased<br />
ADONIS, 6 th March 2009<br />
Are the photosynthetic <strong>response</strong>s to <strong>abiotic</strong><br />
<strong>stress</strong> modulated at transcriptional level?<br />
Annals Botany 2009 103, 609<br />
ADONIS, 6 th March 2009<br />
13
Gene expression <strong>in</strong> rice plants<br />
under drought <strong>stress</strong><br />
Drought <strong>stress</strong> causes<br />
down-regulation of rice<br />
genes cod<strong>in</strong>g for prote<strong>in</strong>s<br />
<strong>in</strong>volved <strong>in</strong> the<br />
photosynthetic light<br />
reactions<br />
Plant Mol Biol 2009 69, 133<br />
ADONIS, 6 th March 2009<br />
TFs <strong>in</strong>volved <strong>in</strong> the photosynthetic<br />
<strong>response</strong> to <strong>abiotic</strong> <strong>stress</strong><br />
Fast <strong>response</strong>s<br />
Long term <strong>response</strong>s<br />
MYB60<br />
Protodermal<br />
Cell<br />
Meristemoid<br />
Mother Cell<br />
Guard<br />
Mother Cell<br />
Mature Guard<br />
Cells<br />
ABI3<br />
MMC<br />
GMC<br />
Stomatal<br />
MYB61<br />
ABA<br />
SPCH<br />
ICE1+SCRM2<br />
ABA<br />
MAP KINASE<br />
PATHWAY<br />
Abiotic <strong>stress</strong><br />
MUTE<br />
ICE1+SCRM2<br />
FLP<br />
MYB88<br />
FAMA<br />
ICE1+SCRM2<br />
Annals Botany 2009 103, 609<br />
ADONIS, 6 th March 2009<br />
14
TFs <strong>in</strong>volved <strong>in</strong> the photosynthetic<br />
<strong>response</strong> to <strong>abiotic</strong> <strong>stress</strong><br />
Abiotic <strong>stress</strong><br />
LIGHT<br />
Non-stomatal<br />
Annals Botany 2009 103, 609<br />
MYB<br />
PSII<br />
PSI<br />
Chloroplast<br />
genes<br />
STZ<br />
AZF2<br />
Glyc<strong>in</strong>e<br />
beta<strong>in</strong>e<br />
Chloroplast<br />
bZIP<br />
Rubisco<br />
Calv<strong>in</strong><br />
cycle<br />
MYB?<br />
σ factor<br />
Mesophyll cell<br />
PpcI and GapI<br />
Nuclear<br />
genes<br />
Cytosol<br />
DOF<br />
PEPCase<br />
Bundle<br />
sheath<br />
ADONIS, 6 th March 2009<br />
NOVEL TRANSCRIPTION FACTORS<br />
REGULATING ABIOTIC STRESS<br />
TOLERANCE IN RICE (<br />
(ORYZA SATIVA L.)<br />
15
Rice <strong>in</strong> Portugal<br />
Consumption: 17 kg/capita/year<br />
Mondego<br />
Production: 60%<br />
Tejo<br />
Sorraia<br />
Sado<br />
PRESENT PROBLEMS:<br />
Pests and diseases<br />
Low production<br />
Infestants<br />
Cold<br />
Sal<strong>in</strong>ity<br />
ADONIS, 6 th March 2009<br />
What makes rice an ideal model<br />
organism?<br />
▪ Important crop<br />
ma<strong>in</strong> source of energy for<br />
2/3 of the world population<br />
▪ Rice genome fully sequenced ~ 400 Mb,<br />
(maize ~ 2500 Mb, barley ~ 5000 Mb, wheat ~ 16000 Mb)<br />
▪ High-efficiency genetic transformation<br />
▪ Genetic and physical maps of high density<br />
▪ High degree of synteny among genes <strong>in</strong> cereal genomes<br />
▪ Insertion knockout mutants available<br />
▪ Microarrays for the whole genome ~50.000 transcripts<br />
RICE - AN IDEAL MODEL ORGANISM FOR<br />
MONOCOTS AND CEREAL CROPS<br />
ADONIS, 6 th March 2009<br />
16
Low temperature signall<strong>in</strong>g pathway<br />
COLD<br />
ICE1<br />
[Ca 2+ ] cyt<br />
U<br />
HOS1<br />
P<br />
ICE1<br />
S<br />
K<strong>in</strong>ases<br />
SIZ1<br />
ICE1-like<br />
U<br />
ICE1<br />
MYB15<br />
Proteolysis<br />
DREB1A/<br />
CBF3<br />
DREB1C/<br />
CBF2<br />
DREB1B/<br />
CBF1<br />
CRT/DRE<br />
COR genes<br />
ACCLIMATION<br />
ADONIS, 6 th March 2009<br />
Yeast one-hybrid screen<strong>in</strong>g to isolate<br />
TFs or other DNA-b<strong>in</strong>d<strong>in</strong>g prote<strong>in</strong>s<br />
2μ ori<br />
pACT II '<br />
PADH1<br />
GAL4-AD<br />
cDNA library<br />
Transformation<br />
Yeast reporter stra<strong>in</strong><br />
(leu2, his3)<br />
LEU2<br />
pINT1 vector<br />
library prote<strong>in</strong><br />
GAL4<br />
AD<br />
<strong>Transcription</strong><br />
if hybrid prote<strong>in</strong><br />
<strong>in</strong>teracts with bait sequence<br />
promoter X<br />
TATA<br />
HIS3 selection gene<br />
Selection for growth on histid<strong>in</strong>e-lack<strong>in</strong>g medium<br />
Ouwerkerk PBF, Meijer AH (2001) Cur. Prot. Mol. Bio.,12.12.1-12.12.22<br />
ADONIS, 6 th March 2009<br />
17
TFs controll<strong>in</strong>g the expression of OsHOS1<br />
ERF1<br />
-1500pb -1000pb -500pb<br />
ERF2<br />
ATG<br />
OsHOS1<br />
0 15´ 30´ 1h 2h 3h 5h<br />
ERF1<br />
Act<strong>in</strong><br />
ERF1 is down-regulated<br />
at low temperature (5ºC)<br />
ERF2 is not regulated at transcriptional level<br />
ADONIS, 6 th March 2009<br />
Low temperature signall<strong>in</strong>g pathway<br />
COLD<br />
ERF1<br />
U<br />
HOS1<br />
P<br />
ICE1<br />
ICE1<br />
S<br />
[Ca 2+ ] cyt<br />
K<strong>in</strong>ases<br />
SIZ1<br />
ICE1-like<br />
U<br />
ICE1<br />
MYB15<br />
Proteolysis<br />
DREB1A/<br />
CBF3<br />
DREB1C/<br />
CBF2<br />
DREB1B/<br />
CBF1<br />
CRT/DRE<br />
COR genes<br />
ACCLIMATION<br />
ADONIS, 6 th March 2009<br />
18
OsDREB1B gene is <strong>in</strong>duced by cold and<br />
drought<br />
Mock<br />
Control<br />
5ºC<br />
Control<br />
10ºC<br />
Control<br />
Drought<br />
Control<br />
0<br />
10’<br />
20’<br />
Shoot<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
0<br />
10’<br />
20’<br />
Root<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
TFs controll<strong>in</strong>g OsDREB1B expression:<br />
GFP::TF<br />
ZF-HD ZF-HD ZF-HD ZF-HD C2H2 bHLH C2H2 C2H2<br />
ATG<br />
OsDREB1B<br />
-1500pb<br />
-1000pb<br />
-500pb<br />
ADONIS, 6 th March 2009<br />
OsSTZ (C2H2) b<strong>in</strong>ds to OsDREB1B promoter<br />
OsSTZ is <strong>in</strong>duced by<br />
cold, salt, and drought<br />
Mock<br />
5ºC<br />
10ºC<br />
ABA<br />
NaCl<br />
Drought<br />
0<br />
10’<br />
20’<br />
Shoot<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
0<br />
10’<br />
20’<br />
Root<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
Mock<br />
5ºC<br />
10ºC<br />
ABA<br />
NaCl<br />
Drought<br />
0<br />
10’<br />
Internal control<br />
20’ 40’ 1h 2h 5h<br />
10h<br />
24h<br />
0<br />
10’<br />
Internal control<br />
20’ 40’ 1h 2h 5h<br />
10h<br />
24h<br />
ADONIS, 6 th March 2009<br />
19
OsPIF (bHLH) b<strong>in</strong>ds to OsDREB1B promoter<br />
The control of OsPIF expression by cold <strong>in</strong>volves<br />
alternative splic<strong>in</strong>g<br />
Mock<br />
0<br />
10’<br />
20’<br />
Shoot<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
0<br />
10’<br />
20’<br />
Root<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
5ºC<br />
10ºC<br />
ABA<br />
NaCl<br />
Drought<br />
Mock<br />
5ºC<br />
10ºC<br />
ABA<br />
NaCl<br />
Drought<br />
0<br />
10’<br />
Internal control<br />
20’ 40’ 1h 2h 5h<br />
10h<br />
24h<br />
0<br />
10’<br />
Internal control<br />
20’ 40’ 1h 2h 5h<br />
10h<br />
24h<br />
ADONIS, 6 th March 2009<br />
A C2H2 that b<strong>in</strong>ds to OsDREB1B promoter<br />
C2H2 transcription is <strong>in</strong>duced at a<br />
higher level <strong>in</strong> the roots<br />
Mock<br />
5ºC<br />
10ºC<br />
ABA<br />
NaCl<br />
Drought<br />
0<br />
10’<br />
20’<br />
Shoot<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
0<br />
10’<br />
20’<br />
Root<br />
40’ 1h 2h<br />
5h<br />
10h<br />
24h<br />
Mock<br />
5ºC<br />
10ºC<br />
ABA<br />
NaCl<br />
Drought<br />
0<br />
10’<br />
Internal control<br />
20’ 40’ 1h 2h 5h<br />
10h<br />
24h<br />
0<br />
10’<br />
Internal control<br />
20’ 40’ 1h 2h 5h<br />
10h<br />
24h<br />
ADONIS, 6 th March 2009<br />
20
Low temperature signall<strong>in</strong>g pathway<br />
COLD<br />
SALINITY<br />
DROUGHT<br />
ERF1<br />
U<br />
HOS1<br />
U<br />
ICE1<br />
ICE1<br />
P<br />
ICE1 S<br />
MYB15<br />
[Ca 2+ ] cyt<br />
K<strong>in</strong>ases<br />
SIZ1<br />
ICE1-like<br />
C2H2<br />
bHLH<br />
STZ<br />
Proteolysis<br />
DREB1A/<br />
CBF3<br />
DREB1C/<br />
CBF2<br />
DREB1B/<br />
CBF1<br />
CRT/DRE<br />
COR genes<br />
ACCLIMATION<br />
ADONIS, 6 th March 2009<br />
OsDREB1A gene is <strong>in</strong>duced by cold<br />
28ºC<br />
5ºC<br />
0<br />
15’<br />
30’<br />
1h<br />
Shoot<br />
2h<br />
3h<br />
5h<br />
7h<br />
12h 24h<br />
OsDREB1A<br />
Control<br />
Control<br />
OsAct<strong>in</strong>1<br />
Control of OsDREB1A expression:<br />
bZIP<br />
ATG<br />
-1500pb<br />
-1000pb<br />
-500pb<br />
OsDREB1A<br />
bZIP<br />
Known to be <strong>in</strong>volved <strong>in</strong><br />
biotic <strong>stress</strong> <strong>response</strong>s<br />
ADONIS, 6 th March 2009<br />
21
Low temperature signall<strong>in</strong>g pathway<br />
COLD<br />
SALINITY<br />
DROUGHT<br />
ERF1<br />
U<br />
HOS1<br />
U<br />
ICE1<br />
ICE1<br />
P<br />
ICE1 S<br />
MYB15<br />
[Ca 2+ ] cyt<br />
K<strong>in</strong>ases<br />
SIZ1<br />
ICE1-like<br />
C2H2<br />
bHLH<br />
STZ<br />
Proteolysis<br />
bZIP<br />
DREB1A/<br />
CBF3<br />
DREB1C/<br />
CBF2<br />
DREB1B/<br />
CBF1<br />
CRT/DRE<br />
COR genes<br />
BIOTIC<br />
ACCLIMATION<br />
ADONIS, 6 th March 2009<br />
Plant cell <strong>response</strong>s to sal<strong>in</strong>ity<br />
SALINITY<br />
High Na +<br />
RMC – Root Meander Curl<strong>in</strong>g<br />
- perception / <strong>response</strong> biotic <strong>stress</strong><br />
RMC<br />
Na<br />
+<br />
H +<br />
Ca +<br />
SOS3<br />
SOS2<br />
P ?<br />
ABI2<br />
H +<br />
SOS2<br />
CBL10<br />
Ca +<br />
Vacuole<br />
CAX1<br />
Ca +<br />
Ca +<br />
CBL10<br />
SOS2<br />
H +<br />
NHX1<br />
Na +<br />
H +<br />
P ?<br />
Li +<br />
SOS3<br />
SOS2<br />
NHX1 -Na<br />
Na + /H + antiport –<br />
tonoplast - <strong>response</strong> to salt <strong>stress</strong><br />
ADONIS, 6 th March 2009<br />
22
The expression of OsRMC and OsNHX1<br />
is <strong>in</strong>duced by NaCl<br />
100mM NaCl<br />
200mM NaCl<br />
OsRMC<br />
0h 2h 5h 12h 24h 2h 5h 12h 24h<br />
OsNHX1<br />
OsAct<strong>in</strong>1<br />
ADONIS, 6 th March 2009<br />
<strong>Transcription</strong>al regulation of OsRMC<br />
-1500pb<br />
ERF3<br />
-1000pb<br />
ERF4<br />
-500pb<br />
ATG<br />
OsRMC<br />
ERF3<br />
Homologue to the Arabidopsis ABR1, a<br />
negative regulator of ABA <strong>response</strong>s<br />
ERF4<br />
Phosphorylated <strong>in</strong> vitro by MPK12/BWMK1<br />
ADONIS, 6 th March 2009<br />
23
Salt signall<strong>in</strong>g pathway<br />
SALINITY<br />
BIOTIC<br />
ERF3<br />
P<br />
ERF4<br />
RMC<br />
Receptor?<br />
TF?<br />
TF?<br />
Responsive Elements<br />
Stress-responsiveresponsive genes<br />
ADONIS, 6 th March 2009<br />
Prelim<strong>in</strong>ary conclusions<br />
▪ The genes analysed are controlled by several TFs,<br />
differentially regulated by different <strong>stress</strong>es<br />
Regulation<br />
<strong>Transcription</strong>al<br />
Post-transcriptional<br />
transcriptional<br />
(and post-translational translational ?)<br />
▪ <strong>Transcription</strong>al regulation of OsDREB1A and OsRMC<br />
cold and sal<strong>in</strong>ity cross talk with<br />
biotic <strong>stress</strong> signall<strong>in</strong>g,<br />
light sens<strong>in</strong>g<br />
ADONIS, 6 th March 2009<br />
24
ACKNOWLEDGEMENTS<br />
Margarida Oliveira<br />
EGP Group<br />
Pieter Ouwerkerk<br />
Leiden University<br />
Duarte Figueiredo (DREB1B)<br />
Tânia Serra (RMC)<br />
Tiago Lourenço (HOS1)<br />
Subhash Chander (DREB1A)<br />
Pedro Barros (cDNA library)<br />
Roberto van Maanen (NHX1)<br />
FCT for the PhD and fellowships,<br />
and the project POCTI/BIA-BCM/56063/2004<br />
ADONIS – Marie Curie Action (tra<strong>in</strong><strong>in</strong>g and mobility)<br />
ADONIS, 6 th March 2009<br />
25
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