signal transduction and gene regulation in plant development

8/13/2019 Signal Transduction and Gene Regulation in Plant Development

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Cellular Signal Transduction


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Signal Transduction and gene

regulation in plant development

and stress responsesSignal transduction is the process by which anextracellular signaling molecule activates amembrane receptor, that in turn alters intracellular

molecules creating a response.There are two stages in this process: a signallingmolecule activates a certain receptor on the cellmembrane, causing a second messenger to

continue the signal into the cell and elicit aphysiological response. In either step, the signalcan be amplified, meaning that one signallingmolecule can cause many responses.


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Signaling molecule

Receptor of target cell

Intracellular molecule

biological effect

Signaltransduction


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Pathway


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Receptor

Receptors are specific membrane

proteins, which are able to recognize

and bind to corresponding ligandmolecules, become activated, and

transduce signal to next signaling

molecules.

Glycoprotein or Lipoprotein


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ligandA small molecule that binds

specifically to a larger one; for

example, a hormone is the ligand for

its specific protein receptor.


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Membrane receptors

membrane

Glycoprotein

Intracellular receptors

Cytosol or nuclei

DNA binding protein


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Signalling molecules

Signal transduction involves the binding of

extracellular signalling molecules and ligands

to cell-surface receptors that trigger events

inside the cell.

Intracellular signaling cascades can be startedthrough cell-substratum interactions.

Receptors- membrane proteins, membrane

potential,proteinaceous pores,channels

C- terminal region of transmembrane protein

receptor is phosphorylated by protein kinases


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Effect bymembrane

receptors

Effect by

intracellular

receptors

Intracellular

molecules

Extracellular

molecules

Signal

molecules

cAMP, cGMP, IP3, DG, Ca2+

Proteins and peptides:

Hormones, cytokinesAmino acid derivatives:

Catecholamines

Fatty acid derivatives:

Prostaglandins

Steroid hormones,Thyroxine, VD3


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Transduction of abiotic signals to altered

gene expression at the cellular level

Ozone,extreme temperatures,flooding, drought,salt

Plants

Physiological & developmental events

Sress recognitionSignal transductionAltered cellularmetabolism


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Gene expression patterns often change

in response to stress:

Changes in metabolism & development leads to alteredgene pattern

These changes are integrated into a response by thewhole plant that may modify growth ,development andeven influence reproductive capabilities

Clues from yeast & bacterial proteins show that theseproteins initiate signal transduction in response toabiotic stress (e.g. low osmotic potential) ,and also

involves hormones (ABA,JA,ethylene) & secondarymessengers (Ca 2+)


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Model depicting various possible

events involved in abiotic stresses

(1)stress perceival,

(2) stress signal

transduction

(3) transcriptional

activation of stress genes(4)synthesis and

accumulation of stress

proteins, resulting finally in

(5) biochemical,(6) cellular and

(7) Physiological

manifestations


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Elements of signal transduction

Intracellular Ca++(Second messenger)-

information from extracellular source to target

with in the cell

Protein kinases (enzyme that phosphorylate &

alter the activity of target protein)-


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Many enzymes are regulated by covalent

attachment of phosphate, in ester linkage, to the

side-chain hydroxyl group of a particular amino acid

residue (serine, threonine, or tyrosine).

H3N+

C COO

CH OH

CH3

H

threonine(Thr)

H3N+

C COO

CH2

OH

H

serine(Ser)


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A protein kinasetransfers the terminal phosphate of

ATP to a hydroxyl group on a protein. A protein phosphatasecatalyzes removal of the Piby

hydrolysis.

Protein OH + ATP Protein O P

O

O

O

+ ADP

Pi H2O

Protein Kinase

Protein Phosphatase


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Phosphorylationmay directly alter activity of an

enzyme, e.g., by promoting a conformational change.

Alternatively, altered activity may result from binding

another proteinthat specifically recognizes a

phosphorylated domain.

E.g., 14-3-3proteins bind to domains that include

phosphorylated Ser or Thrin the sequence

RXXX[pS/pT]XP, where X can be different amino acids.

Binding to 14-3-3is a mechanism by which someproteins (e.g., transcription factors) may be retained

in the cytosol, & prevented from entering the

nucleus.


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Protein kinases and phosphatases are themselves

regulated by complex signal cascades. For example:

Some protein kinases are activated by Ca++

-calmodulin.

Protein Kinase Ais activated by cyclic-AMP(cAMP).

Protein OH + ATP Protein O P

O

O

O

+ ADP

Pi H2O

Protein Kinase

Protein Phosphatase


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Protein Kinase A(cAMP-Dependent Protein Kinase)

transfers Pifrom ATP to OH of a Ser or Thr in a

particular 5-amino acid sequence.

Protein Kinase A in the resting stateis a complex of:

2 catalytic subunits(C)

2 regulatory subunits(R).R2C2


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R2C2

Each regulatory subunit (R) of Protein Kinase A

contains a pseudosubstratesequence, like thesubstrate domain of a target protein but with Ala

substituting for the Ser/Thr.

The pseudosubstrate domain of (R), which lacks ahydroxylthat can be phosphorylated, binds to the

active site of (C), blocking its activity.


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R2C2+ 4 cAMP R2cAMP4+ 2C

When each (R) binds 2 cAMP, a conformational

change causes (R) to release (C).

The catalytic subunits can then catalyzephosphorylation of Ser or Thr on target proteins.

PKIs, Protein Kinase Inhibitors, modulate activity of

the catalytic subunits (C).


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Phosphodiesteraseenzymes

catalyze:

cAMP + H2O AMP

The phosphodiesterase that

cleaves cAMP is activated byphosphorylation catalyzed by

Protein Kinase A.

Thus cAMP stimulates its owndegradation, leading to rapid

turnoff of a cAMP signal.

N

N

N

N

NH2

O

OHO

HH

H

H2C

HO

PO

O-

1'

3'

5' 4'

2'

cAMP


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Signal amplificationis an important feature of signalcascades:

One hormone molecule can lead to formation of

many cAMP molecules.

Each catalytic subunit of Protein Kinase A catalyzes

phosphorylation of many proteins during the life-

time of the cAMP.


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Phosphatidylinositol Signal Cascades

Some hormones activate a signal cascade based on

the membrane lipid phosphatidylinositol.

O P

O

O

H2C

CH

H2C

OCR1

O O C

O

R2

OH

H

OH

H

H

OHH

OH

H

O

H OH

1 6

5

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2

phosphatidyl-inositol


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Kinases sequentially catalyze transfer of Pifrom ATP toOH groups at positions 5 & 4 of the inositol ring, to yield

phosphatidylinositol-4,5-bisphosphate(PIP2).

PIP2is cleaved by the enzyme Phospholipase C.

O P

O

O

H2C

CH

H2C

OCR1

O O C

O

R2

OH

H

OPO32

H

H

OPO32H

OH

H

O

H OH

1 6

5

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2

PIP2

phosphatidylinositol-

4,5-bisphosphate

O


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When a particular GPCR (receptor) is activated, GTP

exchanges for GDP. Gq-GTP activates Phospholipase C.

Ca++, which is required for activity of Phospholipase C,

interacts with () charged residues & with Pimoieties of

the phosphorylated inositol at the active site.

O P

O

O

H2C

CH

H2C

OCR1

O O C

O

R2

OH

H

OPO32

H

H

OPO32H

OH

H

O

H OH

1 6

5

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2

PIP2

phosphatidylinositol-

4,5-bisphosphate

cleavage by

Phospholipase C

Different isoforms

of Phospholipase C

have different

regulatory domains,

& thus respond to

different signals.

A G-protein, Gqactivates one form

of Phospholipase C.


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Cleavage of PIP2, catalyzed by Phospholipase C, yields 2

second messengers:

inositol-1,4,5-trisphosphate(IP3)

diacylglycerol(DG).

Diacylglycerol, with Ca++, activates Protein Kinase C,

which catalyzes phosphorylation of several cellular

proteins, altering their activity.

OHH2C

CH

H2C

OCR1

O O C

O

R2

diacylglycerol

OH

H

OPO32

H

H

OPO32H

OH

H

H OH

OPO32

1 6

5

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2

IP3

inositol-1,4,5-trisphosphate


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IP3activates Ca++-release channels in ER membranes.

Ca++stored in the ER is released to the cytosol, where itmay bind calmodulin, or help activate Protein Kinase C.

Signalturn-offincludes removal of Ca++from the cytosol

via Ca++-ATPase pumps, & degradation of IP3.

Ca++

ATP ADP + Pi

Ca++

IP3

calmodulin

endoplasmicreticulum

Ca++

Ca++-ATPase

Ca -release channel


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Sequential dephosphorylationof IP3by enzyme-catalyzed

hydrolysis yields inositol, a substrate for synthesis of PI.

IP3may instead be phosphorylatedvia specific kinases, to

IP4, IP5or IP6. Some of these have signal roles.

E.g., the IP4inositol-1,3,4,5-tetraphosphate in some cells

stimulates Ca++entry, perhaps by activating plasma

membrane Ca++channels.

OH

H

OH

H

H

OHH

OH

H

H OH

OH

OH

H

OPO32

H

H

OPO32

H

OH

H

H OH

OPO32

(3 steps) +3 Pi

IP3 inositol

O


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The kinasesthat convert PI (phosphatidylinositol) to PIP2

(PI-4,5-P2) transfer Pifrom ATP to OH at positions 4 & 5of the inositol ring.

PI 3-Kinasesinstead catalyze phosphorylation of

phosphatidylinositol at the 3 position of the inositol ring.

O P

O

O

H2C

CH

H2C

OCR1

O O C

O

R2

OH

H

OH

HH

OHH

OPO3

2

H

O

H OH

1 6

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3 4

phosphatidyl-inositol-

3-phosphate

O


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Head-groups of these transiently formed lipids are ligands

for particular pleckstrin homology(PH) & FYVEproteindomainsthat bind proteins to membrane surfaces.

Other protein domains called MARKSare (+) charged, and

their binding to () charged head-groups of lipids like PIP2

is antagonized by Ca++.

O P

O

O

H2C

CH

H2C

OCR1

O O C

O

R2

OH

H

OH

H

H

OHH

OPO32

H

O

H OH

1 6

52

3 4

phosphatidyl-inositol-

3-phosphate

PI-3-P, PI-3,4-P2,

PI-3,4,5-P3, and

PI-4,5-P2havesignaling roles.


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Protein Kinase B(also called Akt) becomes activated when

it is recruited from the cytosol to the plasma membrane

surface by bindingto products of PI-3 Kinase, e.g., PI-3,4,5-P3.

Other kinases at the cytosolic surface of the plasma

membrane then catalyze phosphorylation of Protein

Kinase B, activating it.

Activated Protein Kinase B catalyzes phosphorylationof

Ser or Thr residues of many proteins, with diverse

effects on metabolism, cell growth, and apoptosis.

Downstream metabolic effectsof Protein Kinase B

include stimulation of glycogen synthesis, stimulation

of glycolysis, and inhibition of gluconeogenesis.


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Gene regulation in plant growth

and development

A variety of external & internal signals modify

plant cell metabolism , growth, &

development

The ability of cell s to respond to these signals

is not confined to cells that are still growing

and developing

Mature cells too, can initiate metabolicresponses and can even reinitiate growth and

division in response to signal information

Characteristics of Signal transduction


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Characteristics of Signal transduction

in Plants The stream of signals to which plant cell react

is continuous and complex

Signal transduction uses a net work of

interactions with in the cells, among the cells

and through out the plant

Plant cells contain two information systems;

genetic & epigenetic

Different signals affect the transduction

network in different ways and at different

places, but most modify gene expressson


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Types of signal transduction pathway in

Plants

The bacterial two component system in whicha receptor and an effector interact through

phosphorylation of histidine and aspartate

residueAutophosphorylation of receptor---

Phosphorylation of response regulator---

dephosphorylation of response regulator

(e.g.-Arabdopsis ethylene receptor-ETR1,

&cytokinin sensing CKI1/GCR1)

Plant hormones as a receptor---ethylene,

ABA,GA,IAA,JA,phytochromes etc.


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Saline stress inhibits sucrose synthesis and

promotes accumulation of mannitol

Fructose- 6-Po4

Phosphomannose isomerase

Mannose-6-Po4Mannose -6 Po4 reductase NADPH----NADP

Mannitol-1 po4

Mannitol-1 Po4 phosphatse -Pi

Mannitol


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Accumulation of pinitol in response to

salt stress

Glucose-6 Po4

NAD+ myo- Inositol-1 PO4 synthase

myo-Inositol- 1- PO4

-Pi myo-Inositol- 1- PO4-phosphatsemyo-inositol

S-adenosylmethionine--- myo-inositol-6-O-methyltransferase

S-adenosylhomocysteineOnonitol

Ononitol epimerasePinitol


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SOS signaling pathway for ion homeostasis

under salt stress inArabidopsis.


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Salt stress elicited Ca2+signals are perceived by SOS3, which

activates the protein kinase SOS2.

Activated SOS2 phosphorylates SOS1, a plasma membrane Na+/H+

antiporter, which then transports Na+out of the cytosol.

The transcript level of SOS1 is regulated by the SOS3-SOS2 kinase

complex.

SOS2 also activates the tonoplast Na+/H+antiporter that

sequesters Na+into the vacuole. Na entry into the cytosol through

the Na+transporter HKT1 may also be restricted by SOS2.

ABI1 regulates the gene expression of NHX1, while ABI2 interactswith SOS2 and negatively regulates ion homeostasis either by

inhibiting SOS2 kinase activity or the activities of SOS2 targets.

Double arrow indicates SOS3-independent and SOS2-dependent

pathway.


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ABA-dependent and ABA-

independent signal transduction.


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Environmental factors that increase

concentration of ROS

Ozone- UV radiations +SO2,NO,NO2,--------------Stratasphoric/Troposphoric-

Drought,

Senesence,

Herbicides,(paraquat dichloride)Wounding,

Intense light, --OXIDATIVE STRESS

Pathogens,

Heat&Cold,

Heavy metals,

Root nodules-----------------------------------------


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Generation and scavenging of superoxide radical and

hydrogen peroxide, and hydroxyl radical-induced lipid

peroxidation and glutathione

peroxidase-mediated lipid (fatty acid) stabilization

R l f l i l


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Role of osmolytes in plant

protection against abiotic stress Some compatible solutes may serve protective

functions in addition to OA called osmoprotector.

Glycine betain prevents salt- induced inactivation of

Rubisco & destabilization of the oxygen evolvingcomplex of PSII

Sorbitol,mannitol,myo-inositol, and proline can also

scavenge hydroxyl radicals in addition to OA.

Transgenic plants can be used to test the acclimative

functions of specific osmolytes


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Fatty acids signalling molecules

Protein kinases (PK): primary elements in


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Protein kinases (PK): primary elements in

the signal transduction These are ubiquitous enzymes and are signal specific

Catalyses reversible transfer of-PO4 from ATP toSerine,threonine or tyrosine amino acid chains on target

proteins

Activity is counter balanced by the specific protein

phosphatases Activation of PK has been implicated in response to

light,pathogen attack,GR,salt,heat temp.stress,nutrient

deprivation etc.

Several protein kinases are concerned with regulation ofmetabolic pathways

Hundereds of plant genes fdor different PKs have been

identified but atleast 1000 must exist(Tab,:various groups of

PKs identified in plants

Kinase cascade involved in biotic stress


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Kinase cascade involved in biotic stressresponses in plants

The sensing of stress signals and their transduction into

appropriate responses is crucial for the adaptation andsurvival of plants.

Kinase cascades of the mitogen-activated protein kinase

(MAPK) class play a remarkably important role in plant

signalling of a variety of abiotic and biotic stresses. MAPK

cascade-mediated signalling is an essential step in the

establishment of resistance to pathogens. Here, we describe

the most recent insights into MAPK-mediated pathogen

defence response regulation with a particular focus on the

cascades involving MPK3, MPK4 and MPK6.

We also discuss the strategies developed by plant pathogens

to circumvent, inactivate or even hijack MAPK-mediated

defence responses.


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PAMP- induced MAPK cascade in the plant

defence to the bacterial & fungal

pathogens


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SALT AND DROUGHT STRESS SIGNAL

TRANSDUCTION IN PLANTS

Salt and drought stress signal transduction consists of ionic and osmotic

homeostasis signaling pathways, detoxification (i.e., damage control and

repair) response pathways, and pathways for growth regulation. The ionic

aspect of salt stress is signaled via the SOS pathway where a calcium-

responsive SOS3-SOS2 protein kinase complex controls the expression and

activity of ion transporters such as SOS1. Osmotic stress activates severalprotein kinases including mitogen-activated kinases, which may mediate

osmotic homeostasis and/or detoxification responses. A number of

phospholipid systems are activated by osmotic stress, generating a diverse

array of messenger molecules, some of which may function upstream of

the osmotic stressactivated protein kinases. Abscisic acid biosynthesis isregulated by osmotic stress at multiple steps. Both ABA-dependent and -

independent osmotic stress signaling first modify constitutively expressed

transcription factors, leading to the expression of early response

transcriptional activators, which then activate downstream stress

tolerance effector genes.

I & O ki f


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Inputs & Out puts:making sense fordrought &salt stress signalling

pathways

Figure 1 Functional demarcation of salt and drought stress signaling pathways.

The inputs for ionic and osmotic signaling pathways are ionic (excess NaC) and osmotic (e.g.,

turgor) changes. The output of ionic and osmotic signaling is cellular and plant homeostasis.

Direct input signals for detoxification signaling are derived stresses (i.e., injury), and the signaling

output is damage control and repair (e.g., activation of dehydration tolerance genes). Interactions

between the homeostasis, growth regulation, and detoxification pathways are indicated


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THE SOS REGULATORY PATHWAY FOR ION

HOMEOSTASIS AND SALT TOLERANCE

High NaC stress initiates a calcium

signal that activates the SOS3-

SOS2 protein kinase complex,

which then stimulates the NaC/HC

exchange activity of SOS1 and

regulates transcriptionally

and posttranscriptionally theexpression of some genes. SOS3-

SOS2 may also stimulate or

suppress the activities of other

transporters involved in ion

homeostasis under salt stress,

such as vacuolar HC-ATPases and

pyrophosphatases (PPase),vacuolar NaC/HC exchanger

(NHX), and plasma membrane KC

and NaC transporters.

Figure 2 Regulation of ion (e.g., NaC and KC) homeostasis by

the SOS pathway.


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PROTEIN KINASE PATHWAYS FOR

OSMOTIC STRESS SIGNALING

Figure 3 Activation of protein kinases by hyperosmotic stress.

The MAP kinase cascade shown is also activated by other stresses.

Currently, the functional significance of the kinase activation is unclear

(hence the unknownoutput). SIPK, SIMK, and ATMPK6 are homologous

MAP kinases from tobacco, alfalfa, and Arabidopsis, respectively.


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OSMOTIC STRESSACTIVATED

PHOSPHOLIPID SIGNALING

Figure 4 Phospholipid signalingunder salt stress, drought, cold, or

ABA.

Osmotic stress,cold, and ABA activate

several types of phospholipases that

cleave phospholipids to generate lipid

messengers (e.g., PA, DAG, and IP3),which regulate stress tolerance partly

through modulation of stress-

responsive gene expression.FRY1(a 1-

phosphatase) and 5-

phosphatasemediated

IP3 degradation attenuates the stressgene regulation by helping to control

cellular IP3 levels. PLC, phospholipase

C; PLD, phospholipase D; PLA2,

phospholipase A2; PA, phosphatidic

acid; DAG, diacyglycerol.

S d l d G


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Stress- and ABA-Regulated Gene

Expression

Figure 5 ABA

metabolism is regulated

by osmotic stress at

multiple steps.

The ABA biosynthesis

genes ZEP, NCED,

LOS5/ABA3, and AAO are

upregulated by salt and

drought stresses. ABA

degradation is also

important in controlling

cellular ABA content, and

biochemical evidence

suggests osmotic stress

inhibition of the first step

of catabolism

ABA Dependent and ABA


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ABA-Dependent and ABA-

Independent Signaling

Figure 6 Model showing osmotic

stress regulation of early-

response and delayedresponse

genes. (A) Model integrating stress

sensing, activation of phospholipid

signaling and MAP kinase cascade,

and transcription cascade leading to

the expression of delayed-response

genes. (B) Examples of early-response

genes encoding inducible transcription

activators and their downstreamdelayed-response genes encoding

stress tolerance effector proteins.

Question marks denote unknown

transcription factors that activate the

early-response genes.

F tt id d i d i l i l t


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Fatty acid - derived signals in plants

e.g. Jasmonates

Its mutant opr3 in biosynthetic pathway,

allow the dissection of cyclopentanone &

cyclopentenone signalling

In addition, keto,hydroxy & hyperhydroxy FA

involved in cell death & stress related gene

expression.

Bruchins & volicitin as a signal molecule frominsects show FA 0derived signalling in plant

defence

signal transduction and gene regulation in plant development

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