TEMA 4.

RUTAS DE TRANSDUCCIÓN DE SEÑALES EN EL SISTEMA

DE DEFENSA DE LAS PLANTAS

 REGULACIÓN HORMONAL

(SA, JA, ET, ABA, y otras rutas de transducción).

 ESPECIES REACTIVAS DE OXÍGENO Y

NITRÓGENO
 DEFENCE SIGNALLING NETWORKS IN ARABIDOPSIS
PHYTOPATHOGENIC
BACTERIA AND FUNGI

R pad4
eds1
ndr1
JA coi1 E pad3
eds5
cev1 jar1 ein2 coi1 SA
PATHWAY fad4 sid2
cpr5 ctr1 etr1 cpr1
npr1 NahG
cet ein3 edr1
mpk4 cpr5 npr1
mpk4 cpr6
dth9
DEFENCE TH2.1 PDF1.2 Camalexin PR1a
MOLECULES Phytoalexins PR4(Hel) PR2
PR1b PR5
PR3(b-Chi)

RESISTANCE Pythium sp. Erysiphe sp. , B. cinerea, P. porri P. syringae, P. parasitica,

AGAINST A. brassicicola E. carotovora, Erysiphe sp. , X. campestris
B. cinerea O. lycopersicum E. carotovora,
DEFENCE SIGNALLING NETWORKS IN ARABIDOPSIS: SA

Nandi et al., 2004

LOSS OF SA ACCUMMULATION CONFERS SUSCEPTIBILITY

The bacterial enzyme encoded by NahG

hydrolyzes SA to catechol

Salicylate
Before SA-deficient
hydroxylase
plants were identified
encoded by
genetically, NahG
NahG
expression was used to
investigate SA’s roles.
SA catechol
NahG-expressing
Control
plant Although it is very effective,
concerns about its specificity
and effects of catechol mean
researchers now prefer other
methods

From Delaney, T.P., Uknes, S., Vernooij, B., Friedrich, L., Weymann, K., Negrotto, D., Gaffney, T., Gut-Rella, M., Kessmann, H., Ward, E. and Ryals, J. (1994). A central role of salicylic acid in plant
disease resistance. Science. 266: 1247-1250, reprinted with permission from AAAS.
SA BIOSYNTHESIS

=SID2

isochorismate
pyruvate lyase

Wildermuth et al., 2001; Dempsey et al., 2011

SA BIOSYNTHESIS Expression of ICS1
increases upon
exposure to a pathogen

PR1 PR1

ICS1

ICS1

sid 2-2/eds16-1

sid 2-1

sid2: sa induction-deficient Wildermuth et al., 2001

 SA BIOSYNTHESIS

SA is synthesized in the chloroplast, conjugated in

the cytoplasm and sequestered in the vacuole

plastid

SA
ICS

-glucose
SA
SAG SAG (Salicylic acid O-β-glucoside)
SGE

MeSA
-glucose

SGE (Salicyloyl glucose ester)

 SA BIOSYNTHESIS

O O-
OH

Dihydroxybenzoates
SA
2-Hydroxybenzoic Acid hydroxylases
SA

PBS3
UDP- glucosyltransferase SABP2 SA aa synthase
Methyl esterase
SA carboxil methyltransferase
SAMT

SAG SA-aa
vacuoles MeSA
Loake and Grant, 2007; Zhang et al., 2017
Zhang et al., 2017
 SA BIOSYNTHESIS

Mutants in ICS1 are blocked in SA synthesis and

susceptible to disease
Col-0
Reduced
accumulation of free
and conjugated SA

sid2 and eds16 are

ICS1 mutants
sid2

Increased growth of
Hyaloperonospora
arabidopsidis
pathogen

Nawrath, C. and Metraux, J.-P. (1999). Salicylic acid induction–deficient mutants of Arabidopsis express PR-2 and PR-5 and accumulate high levels of camalexin
after pathogen inoculation. Plant Cell. 11: 1393-1404.
 KEY COMPONENTS OF
THE SA PATHWAY

MUTANTS ISOLATION

SA receptor WANTED!!

Seyfferth y Tsuda, 2014

 KEY COMPONENTS OF THE SA PATHWAY

The eds1 mutant shows enhanced disease

susceptibility

Wild type:
eds1:
Pathogen
Plant fails to
growth is
respond and
arrested by
pathogen
plant Wild-type eds1
growth is
defense
unchecked
response

Parker, J.E., Holub, E.B., Frost, L.N., Falk, A., Gunn, N.D., and Daniels, M.J. (1996). Characterization of eds1, a mutation in Arabidopsis suppressing resistance to Peronospora
parasitica specified by several different RPP genes. Plant Cell 8: 2033-2046.
 KEY COMPONENTS OF THE SA PATHWAY

The eds1 mutant is deficient in PR1

gene expression but rescued by SA

Pathogen
recognition

Wild-type
EDS1

eds1 SA
SA

No PR1
induction by Defense
pathogen response

Falk, A., et al., 1999; Cui et al., 2017

 KEY COMPONENTS OF THE SA PATHWAY

npr1/nim1 has affected SA signaling

Virulent Pto

Cao et al., 1994

 KEY COMPONENTS OF THE SA PATHWAY

SA plays a key role amplifying the defence signallig and also

its accumulation contributes to a REDUCING environment

NPR1 oligomerizes via

Monomeric NPR1 is
redox-sensitive Cysteines imported into the nucleus
(here it is GFP-labeled)
SA

TGAs

TA TA A AT G TA A
ATATT TAC ATT

NPR-1 Wu et al., 2012

CYTOSOL DEFENCE GENES
 KEY COMPONENTS OF THE SA PATHWAY

 NPR1 does not bind directly to DNA

 NPR1 is part of a multigene family (NPR3, NPR4 also
interact with SA)
 Two alternative but not exclusive models for SA
perception are proposed.
 NPR1 is the SA receptor
 NPR3 and NPR4 perceive SA regulating
NPR1 protein accumulation

Wu et al., 2012; Fu et al., 2012, Seyfferth and Tsuda, 2014

 ROLE OF NPR1

NPR1 Kd=140nM
Inactive
SA Active NPR3 Kd=981nM

NPR4 Kd= 46nM

De-repression of
transactivation domain

Seyfferth y Tsuda, 2014

 NPR1 MODE OF ACTION

Seyfferth y Tsuda, 2014 Wang et al., 2006

DEFENCE SIGNALLING NETWORK IN ARABIDOPSIS: SA

WRKYs are a large family of transcription

factors
There are 74 Arabidopsis WRKYs and 90 rice WRKYs, many of which are
involved in defense signaling. Some promote and some repress transcription

Genes induced by WRKYs

-WRKY
include NPR1 and ICS1 →
signal amplification!
+WRKY

NPR1
W- box ICS1
SA NPR1 +WRKY
C/TTGACT/C
IDENTIFICATION OF MORE SA-MEDIATED PATHWAY COMPONENTS

In the sni1 mutant, PR-1 genes are SNI1 maintains a low

expressed all the time even in level of expression in
non-inducing conditions non-inducing conditions
In the absence of SA:
Gene OFF

control
SNI1

pathogen In the presence of SA:

Gene ON

npr1 = no PR expression
npr1sni1 = suppressor of npr1 = constitutive PR
expression
SNI1 inhibits activating
chromatin remodelling
Li et al., 1999; Song et al., 2011; Kim et al., 2012
DEFENCE SIGNALLING NETWORK IN ARABIDOPSIS: SA

Durrant et al., 2007; Yan et al., 2013

 JASMONATE SIGNALLING

JA and related molecules

REGULATE THE FOLLOWING RESPONSES

maduración del fruto

patógenos
crecimiento de la raíz
heridas
tuberización
estrés hídrico
desarrollo del polen
exposición a ozono
JA BIOSYNTHESIS 18:3 (Δ9,12,15) LIPASES

Allene oxide synthase

chloroplast Allene oxide cyclase

12 oxo-phytodienoic acid reductase 3

peroxisome
Acyl-CoA oxidase

JASMONATES

* JMT

JAR1
*
Balbi and Devoto, 2008
JASMONATE SIGNALlING

Highly active enantiomer Staswick, 2008

JASMONIC ACID SIGNALLING PATHWAY

Pieterse et al., 2012

 JASMONIC ACID SIGNALLING PATHWAY

THE JAZ FAMILY

Staswick, 2008
Gimenez-Ibañez et al., 2017
 ETHYLENE DEFENCE PATHWAY

JA ET JA

MYC branch

ERF branch
ETHYLENE SIGNAL TRANSDUCTION PATHWAY

Inactivos (ER)

inactiva

Chen et al., 2005

Binder et al., 2007
Merchante et al., 2017
ETHYLENE SIGNALLING
Last step is ERF1 activation
WT 35S:: ERF1.1

Berrocal-Lobo et al., 2002

 ETHYLENE SIGNAL TRANSDUCTION PATHWAY

Constitutive expression of ERF1 increases Arabidopsis

susceptibility to Pseudomonas syringae pv. tomato DC3000

8 W WT
T
N aNahG
hG
W T /ER F 1.1
7 W 35S::ERF1.1
T /ER F 1.2
W 35S::ERF1.2
T /ER F 1.3
35S::ERF1.3
6
log(cfu/mg)

1
0 2 4
I
DAYS
 OTHER SIGNALLING PATHWAYS

 CELL WALL-DERIVED SIGNALLING

 ABCISIC ACID PATHWAY (ABA)

 AUXINS DEFENCE SIGNALLING

 DAMPs RECOGNITION

 OLIGOGALACTURONIDES (OGs)

OGs- INDUCED RESISTANCE IS SA, JA and ET INDEPENDENT

Ferrari et al., 2007

OTHER SIGNALLING PATHWAYS: CELL WALL-DERIVED SIGNALLING

Apoplast

AtCESA4
AtCESA7 AtCESA8
CESA Complex
AtCESA8 AtCESA7
Plasma Membrane
AtCESA4
Cytoplasm

Gene AtCESA Mutant

At4g18780 AtCesA8 ern1/irx1/lew2
AtCESA of Mutants irx
Secondary At5g17420 AtCesA7 irx3 (irregular
cell wall At5g44030 AtCesA4 nws2/irx5
xylem)

At5g05170 AtCesA3 ixr1/cev1 Mutants ixr

AtCESA of
Primary At5g64740 AtCesA6 ixr2 (isoxaben resistant)
cell wall At4g32410 AtCesA1 rsw1
OTHER SIGNALLING PATHWAYS: CELL WALL-DERIVED SIGNALLING

HIGHLY SPECIFIC DEFENCE RESPONSE

PRIMARY CELL WALL MUTANTS SECONDARY CELL WALL MUTANTS

Hernández-Blanco et al., 2007

OTHER SIGNALLING PATHWAYS: CELL WALL-DERIVED SIGNALLING

irx1- MEDIATED RESISTANCE IS SA, JA and ET INDEPENDENT

Hernández-Blanco et al., 2007

OTHER SIGNALLING PATHWAYS: CELL WALL-DERIVED SIGNALLING

CELL WALL MODIFICATIONS ACTIVATE ABA-DEPENDENT

DEFENCE RESPONSES

Hernández-Blanco et al., 2007

ABA DORMANCY

GERMINATION

DEVELOPMENT

STOMATA OPENING/CLOSURE

BIOTIC STRESS RESPONSE

 OTHER SIGNALLING PATHWAYS: ABA
Triple snrk2 mutant
(snrk2.2, 2,3, 2.6)
ABA DORMANCY

GERMINATION

DEVELOPMENT

STOMATA OPENING/CLOSURE

ABA
BIOTIC STRESS RESPONSE
OTHER SIGNALLING PATHWAYS: ABA
Zeaxanthin is
abundant in green
tissues but can be
limiting for ABA
synthesis in roots
OTHER SIGNALLING PATHWAYS: ABA
ABA levels are also controlled
by inactivation pathways ABA is irreversibly
deactivated by conversion
to phaseic acid

Rehydration
Developmental signals

[ABA]
ABA can be reversibly
inactivated by
glucosylation
OTHER SIGNALLING PATHWAYS: ABA

ABA PERCEPTION

INTRACELLULAR EXTRACELLULAR

PUTATIVE RECEPTORS:

1. FCA, RNA-binding protein (Nature, 2006/ Risk et al., 2008)

2. CHLH, chloroplastic protein (Shen et al., 2006)
3. GCR2, 7TM protein (Liu et al, 2007.Science/ Gao et al.,2007)
4. 2 GPCR, GTG1 and GTG2 (Pandey et al., 2009. Cell)

The ABA receptors-we report you decide. McCourt and Creelman, 2008
OTHER SIGNALLING PATHWAYS: ABA

The PYR/RCAR ABA receptors made Science

magazines Top 10 list

GFP:RCAR1

PYR/RCAR
proteins are found
in the cytoplasm
and nucleus (arrow)
RCAR1pro:GFP

From Ma, Y., Szostkiewicz, I., Korte, A., Moes, D.l., Yang, Y., Christmann, A., and Grill, E. (2009). Regulators of PP2C phosphatase activity function as abscisic acid
sensors. Science 324: 1064-1068 reprinted with permission from AAAS.
OTHER SIGNALLING PATHWAYS: ABA

Perception and Signaling

ABA The 14 PYR/RCARs in Arabidopsis

PYR1 PYR/RCAR receptors

Phosphatase PP2C Protein phosphatases

(including ABI1)

Protein
kinases Kinase
(including
SnRK2s and
CDPKs) P
P
TF
ABA RESPONSES
 OTHER SIGNALLING PATHWAYS: ABA

PYR/RCAR receptors bind ABA in a complex with ABI1

or other PP2Cs

NO ABA ABA

PYR1 PYR1

PP2C

PP2C

Reprinted from Raghavendra, A.S., Gonugunta, V.K., Christmann, A., and Grill, E. (2010) ABA
perception and signalling. Trends Plant Sci. 15: 395-401 with permission from Elsevier.
 OTHER SIGNALLING PATHWAYS: ABA

abi1-1 mutants are ABA-insensitive in all their

responses
Germination is
not inhibited on Root growth is
ABA not inhibited on
ABA

ABI1 encodes
a PP2C protein
phosphatase

Guard cells are

not ABA-
Wild responsive
abi1
type
 OTHER SIGNALLING PATHWAYS: ABA

The abi1 mutation stabilizes the inhibitory

effect of ABI1
WILD TYPE WILD TYPE + ABA abi1-1 + ABA
PYR1
PYR1 PYR1

ABI1
abi1-1
ABI1

SnRK2 Kinase
SnRK2
SnRK2
P
P P P
NO ABA ABA NO ABA
RESPONSES RESPONSES RESPONSES
 OTHER SIGNALLING PATHWAYS: ABA

irx1 MUTANT IS RESISTANT AND ACCUMULATES ABA

WHAT HAPPEN IF WE ANALIZED ABA MUTANTS?
 OTHER SIGNALLING PATHWAYS: ABA

irx1 MUTANT IS RESISTANT AND ACCUMULATES ABA

WHAT HAPPEN IF WE ANALIZE ABA MUTANTS?

ABA promotes RESISTANCE in some plant-pathogen interactions,

Whereas it INCREASES SUSCEPTIBILITY in others (Ton et al., 2009)
OTHER SIGNALLING PATHWAYS: ABA

Hernández-Blanco et al., 2007

 OTHER SIGNALLING PATHWAYS: ABA

ABA has both positive and negative effects on disease

resistance

ABA

Defense
response
This pathogen is more
virulent in ABA mutants, This pathogen induces ABA
meaning ABA enhances synthesis which makes the
resistance. plant less resistant.
Pseudomonas syringae is a
Pythium irregulare is a necrotrophic
biotrophic pathogen.
pathogen.
Adie, B.A.T., Perez-Perez, J., Perez-Perez, M.M., Godoy, M., Sanchez-Serrano, J.-J., Schmelz, E.A., and Solano, R. (2007). ABA is an essential signal for plant
resistance to pathogens affecting JA biosynthesis and the activation of defenses in Arabidopsis. Plant Cell 19: 1665-1681; Reprinted by permission from Macmillan
Publishers Ltd. de Torres-Zabala, M., Truman, W., Bennett, M.H., Lafforgue, G., Mansfield, J.W., Rodriguez Egea, P., Bogre, L., and Grant, M. (2007).
Pseudomonas syringae pv. tomato hijacks the Arabidopsis abscisic acid signalling pathway to cause disease. EMBO J 26: 1434-1443 copyright 2007.
 ABA ROLE IN DISEASE RESISTANCE

ABA enhances some

biotic defense
responses and
interferes with others

ITS ROLE IN DISEASE RESISTANCE

DEPENDS ON THE TYPE OF PATHOGEN Ton et al., 2009
 Bacteria
move
towards
open
stomata

Closed
Water
stoma
Open Bacteria
stoma
Pathogens
can produce
Some pathogens override the compounds
plant’s response and reopen that override
stomata the ABA
response

Coronatine is a pathogen-
produced compound
interferes with ABA-induced
stomatal closure.
Yi, H., Preuss, M.L., and Jez, J.M. (2009). The devil (and an active jasmonate hormone) is in the details. Nat Chem Biol 5: 273-274. Melotto, M., Underwood, W., Koczan, J., Nomura,
K., and He, S.Y. (2006). Plant stomata function in innate immunity against bacterial invasion. Cell 126: 969-980.
 OTHER SIGNALLING PATHWAYS: AUXINS

AUXINS
AUXIN SA/BTH
flg22

DEGRADATION OF REGULATORS

RESISTANCE TO RESISTANCE TO
NECROTROPHS BIOTROPHS

Llorente et al., 2008 Navarro et al., 2006

The auxin-receptor TIR1 is a component of
the SCF ubiquitin ligase complex SCFTIR1

F-box TIR1
auxin
receptor F-box TIR1
protein
SKP1

SCF complex

CUL1
SCF ubiquitin ligase complex
(Named for SKP1, CUL1 and F-box
proteins)
Aux/IAA proteins repress the activity of
Auxin Response Factors (ARFs)

ARF and Aux/IAA proteins have ARF

DNA- Activation / III IV
sequence similarity in their C- binding repression
terminal regions through which domain domain
they can homodimerize.
Aux/IAA
I II III IV

ARF homodimer

ARF Aux/IAA heterodimer

 The auxin signaling pathway is
amazingly short!

Auxin OUTPUT

INPUT
 Perhaps the diversity of auxin responses
is correlated with the diversity in signaling
proteins
Arabidopsis has 29 Arabidopsis has 23
Aux/IAA proteins ARF proteins

Auxin OUTPUT

INPUT
The number of potential
interactions between
ARFs and Aux/IAA
proteins is quite large,
and has the potential to
TIR1 is related to precisely control auxin-
five other auxin- induced gene
receptor proteins, expression in a vast
AFB1 – AFB5. number of
combinations.
CROSSTALK BETWEEN THE DIFERENT SIGNALLING PATHWAYS
 SA AND JA CROSSTALK

Infection with biotrophic Pseudomonas syringae,

which induces SA-mediated defense,
rendered plants more susceptible to the necrotrophic
pathogen Alternaria brassicicola by
suppression of the JA signaling pathway.

Spoel et al., 2007

SA AND JA CROSSTALK

Spoel et al., 2007

 MPK4

PAD4 EDS1

Adapted from Pieterse et al., 2012

CROSSTALK BETWEEN THE DIFERENT SIGNALLING PATHWAYS

ET JA
+ JA JAZ is degraded

+ JA + SA JAZ is degraded
SA ORA 59
via NPR-1
and TGA2, 5, 6
JAZ

PDF1.2

TGA TF ?
EIN3

ORA 59
CROSSTALK BETWEEN THE DIFERENT SIGNALLING PATHWAYS

Pathogen
Elicitors
[SA] [JA] [C2H4]

ERF1
NPR1

SA-responsive genes JA and C2H4 responsive genes

Defense responses