ABIOTIC STRESSES REDIRECT PROTEIN SYNTHESIS Mikal E. Saltveit Mann Laboratory, Department of...

ABIOTIC STRESSESABIOTIC STRESSES REDIRECT PROTEIN REDIRECT PROTEIN

SYNTHESISSYNTHESIS

Mikal E. SaltveitMann Laboratory, Department of Vegetable Crops

University of California, Davis

Participants in this research

M.E. Mangrich J.G. Loaiza-Velarde

F.A. Tomás-Barberán M.A. Ritenour

A. RabM. Cantwell

G. Peiser G. López-Gálvez

A A Stimulus ProducesStimulus Produces A Physiological A Physiological

ResponseResponse

Cause and Effect Occur in a Linear Order!

The StimulusWoundingWounding

Produces the Physiological ResponseIncreased PAL ActivityIncreased PAL Activity

Resulting inTissue BrowningTissue Browning

Lettuce leaf tissue

CUT

Signal Signal

CUT

PAL PAL

Lettuce leaf tissue

Wound

Signal

Receptor

DNA

mRNA

Proteinsynthesis

PALPhenylpropanoid metabolism

Receptor

DNA

mRNA

Proteinsynthesis

PAL-IF

Preexistingphenolics

Vacuole

PPO

PODCellularmembranes

PhenolicsPhenolics

Tissuebrowning

Increasedpermeability

0

20

40

60

80

100

Time

Re

lativ

e v

alu

es

Wound signal PAL activity

PAL inactivating factor

Phenolic compounds

HO-

C

OHCOOH

HO

HO

O

O

p-Coumaroylquinic acid

HO-

C

OHCOOH

HO

HO

O

Feruloylquinic acid

CH O-

3

O

OH

COOH

HO

HO

O

4-Caffeoylquinic acidHO-

C

OHCOOH

HO

HO

O

3-Caffeoylquinic acid

HO-

O

3,5-Dicaffeoylquinic acid3,4-Dicaffeoylquinic acid

Quinic acid

OH

HO

COOHHO

HO

NH2

COOHHO-

COOH

HO-

HO-

Phenylalanine Cinnamate p-Coumarate Caffeate Ferulate

HO-

CH O-

3

4,5-Dicaffeoylquinic acid

5-Caffeoylquinic acid(Chlorogenic acid)

COOH COOH COOH

HO-

C

HO-

O

HO-

C

OHCOOH

HO

O

HO-

O

O

HO-

C

HO-

O

HO-

C

OHCOOH

HO

HO

OHO-

O

(Isochlorogenic acid)

HO-

C

OHCOOH

HO

OHO-

O

HO-

CO

HO-

OHO-

C

OHCOOH

HO

OHO-

O

O

HO-

C

HO-

O

COOH

C

C

COOH

O

-

Dicaffeoyltartaric acid

COOH

C

C

COOH

OH--O

HO-

HO-C-

O

Caffeoyltartaric acid

-OH

-OH

C

OCOOH

C

C

COOH

OH--HO

Tartaric acid

O

HO-

HO-

C-O

Caf

feo

ylta

rtar

ic a

cid

12

0

Butter leafC

hlor

ogen

ic a

cid

60

50

40

30

20

10

0

LSD 5%

Iceberg Romaine

24 48Hours

6 12 720 24 48Hours

6 12 72

50

40

30

20

10

0Dic

affe

oylta

rtar

ic a

cid

0 24 48Hours

6 12 72

Isoc

hlor

ogen

ic a

cid

24

20

16

12

8

4

0

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

LSD 5%

0

Wound, 10 °C

Wound, 5 °C

Ethylene, 10 °C

Ethylene, 5 °C

16

8

4

Iceberg lettuce

Heat-shock

Cut and washed

Held at 5 °C in 95% RH

Periodically assayed for PAL and phenolic compounds

20 40 60

Abs

orba

nce

at 3

20 n

m/g

0.28

0.24

0.20

0.16

0.12

0.08

0.04

0.00

60 sec

30 50 70

Temperature (°C)

How doesheat-shock

reduce subsequent browning in lettuce?

0 200 400 600

Seconds of heat-shock

0.14

0.12

0.10

0.08

0.06

0.04

0.02

0.00100 300 500

PA

L ac

tivity

(µ

mo

l·g

h

)-1-1

45 °C

50 °C

55 °C

Wounded (24 h)

Control (0 h)

120 sec

90 sec

60 sec

~ 6x

Hours

'L' V

alue

0 24 48 72

72

71

70

69

68

67

66

Control

50 °C, 90 sec

55 °C, 60 sec

'a' V

alue

-1

-2

-3

-4

-5

50 °C, 90 sec

55 °C, 60 sec

45 °C, 120 sec

45 °C, 120 sec

Control

Hours0 24 48 72

80

70

60

50

40

30

20

10

0

Control

45 °C, 120 sec

50 °C, 90 sec

55 °C, 60 sec55 °C, 60 sec

Tot

al p

heno

lic c

onte

nt (

µg·

g )-1

HO-

C

OHCOOH

HO

HO

O

O

p-Coumaroylquinic acid

HO-

C

OHCOOH

HO

HO

O

Feruloylquinic acid

CH O-

3

O

OH

COOH

HO

HO

O

4-Caffeoylquinic acidHO-

C

OHCOOH

HO

HO

O

3-Caffeoylquinic acid

HO-

O

3,5-Dicaffeoylquinic acid3,4-Dicaffeoylquinic acid

Quinic acid

OH

HO

COOHHO

HO

NH2

COOHHO-

COOH

HO-

HO-

Phenylalanine Cinnamate p-Coumarate Caffeate Ferulate

HO-

CH O-

3

4,5-Dicaffeoylquinic acid

5-Caffeoylquinic acid(Chlorogenic acid)

COOH COOH COOH

HO-

C

HO-

O

HO-

C

OHCOOH

HO

O

HO-

O

O

HO-

C

HO-

O

HO-

C

OHCOOH

HO

HO

OHO-

O

(Isochlorogenic acid)

HO-

C

OHCOOH

HO

OHO-

O

HO-

CO

HO-

OHO-

C

OHCOOH

HO

OHO-

O

O

HO-

C

HO-

O

COOH

C

C

COOH

O

-

Dicaffeoyltartaric acid

COOH

C

C

COOH

OH--O

HO-

HO-C-

O

Caffeoyltartaric acid

-OH

-OH

C

OCOOH

C

C

COOH

OH--HO

Tartaric acid

O

HO-

HO-

C-O

55 °C, 60 sec

8

6

4

2

0

Control

45 °C, 120 sec 50 °C, 90 sec

55 °C, 60 sec

Hours0 24 48 72

24

20

16

12

8

4

0

Control

45 °C, 120 sec

50 °C, 90 sec55 °C, 60 sec

Caffeoyl tartaric acid

Dicaffeoyl tartaric acid

LSD 5% =

LSD 5% =

Phe

nolic

aci

ds (

µg·

g )

10-1

24

20

16

12

8

4

0

Control 45 °C, 120 sec

50 °C, 90 sec

55 °C, 60 sec

10

8

6

4

2

Control

45 °C, 120 sec

50 °C, 90 sec

55 °C, 60 sec

5-Caffeoyl quinic acid (Chlorogenic acid)

3,5-Dicaffeoyl quinic acid (Isochlorogenic acid)

LSD 5% =

LSD 5% =

Phe

nolic

aci

ds (

µg·

g )

Hours0 24 48 72

-1

Application of a Application of a heat-shock heat-shock after after

woundingwounding

0

0.1

0.2

0.3

0.4

0 5 10 15 20 25 30 35 40

Hours after wounding

PA

L ac

tivity

ControlControl

HS after 8 hHS after 8 h

HS after 1 hHS after 1 h

HS after 24 h

Application of a Application of a heat-shock heat-shock

beforebefore wounding wounding

WoundedWoundedcontrolcontrol

HS -4 h, WHS -4 h, W

W, HS +2 h

W, HS +2 h, W +6 h

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

0 5 10 15 20 25 30 35 40

Hours after wounding

PA

L ac

tivity

0.00

0.02

0.04

0.06

0.08

0.10

0.12

-40 -35 -30 -25 -20 -15 -10 -5 0 5

Wounded

Unwoundedcontrol

Heat-shocked

Hours before wounding

PA

L ac

tivity

Wounded

50 °C, 90 sec

Held at 5 °C

It’s not the proteins that are

synthesizedbut the

synthesis of the proteinsthat

protects against browning

Protein synthesis

““NormaNormal”l”

GeneralGeneral

Protein synthesis

““Normal”Normal”

GeneralGeneral Wounding

SignalSignal

PAL

Heat-shock proteins

Heat-shock

Protein synthesis


GeneralGeneral

“Signal”

Could the linear sequence of metabolic events induced by one stress (e.g., wounding)

be redirected by another abiotic stress

(e.g., heat-shock)?

PALHeat-shock proteins

Heat-shock

Protein synthesis


Wounding

SignalSignal“Signal”

GeneralGeneral

Heat-shock

Wounding (e.g., of lettuce)

PAL

Phenolics accumulate

Few phenolic compounds present

Very little browningVery little browning

Normal conditions

X

Browning

SignaSignall

No wound No wound signal remainingsignal remaining

Synthesis of HSPs

Recovery from the heat-shock and resumption of normal protein synthesis

It appears that one stress It appears that one stress

((heat shockheat shock) can redirect ) can redirect

and modify the response of and modify the response of

another stress (another stress (woundingwounding))

Chilling Injury is Chilling Injury is Another Stress that is Another Stress that is

Thought to Occur Thought to Occur Through a Linear Through a Linear

Sequence of EventsSequence of Events

Chilling temperaturesChilling temperatures

Membrane phase changeMembrane phase change

Secondary effectsSecondary effects

Primary effectPrimary effect

Primary cause

Increased membrane permeability Influx of calciumDepolymerization of microtubules Reduced photosynthesisProtoplasmic streaming stops Altered enzyme activityToxic metabolites accumulate Altered metabolism

SymptomsSymptomsAccelerated senescence Increased decay Increased water loss Vascular browningAbnormal ripening Tissue discolorationIncreased ethylene production Elevated respiration

Does Their Synthesis Prevent Does Their Synthesis Prevent the Synthesis of an Agent the Synthesis of an Agent Causing Chilling InjuryCausing Chilling Injury??

Do Heat-shock Proteins Protect Do Heat-shock Proteins Protect Against Chilling Injury?Against Chilling Injury?

or

Chilling temperatures

Abnormal metabolism

Accumulated toxins

Chilling injury

Heat-shock

HSPs


Abnormal metabolism

Accumulated toxins

Heat-shock

HSPs

Chilling injuryNo chilling injury


Abnormal metabolism

Accumulated toxins

Chilling injury

Heat-shock


Abnormal metabolism

Accumulated toxins

Chilling injury

Heat-shock

HSPs

No accumulated toxins

No chilling injury

Chilling temperaturesChilling temperatures

Primary effectsPrimary effects

Membranes Microtubules Proteins Pathways

Primary effectsPrimary effects Primary effectsPrimary effects

Secondary effectsSecondary effects Secondary effectsSecondary effectsIncreased membrane permeability Influx of calciumDepolymerization of microtubules Protoplasmic streaming stopsAltered enzyme activity Altered metabolismReduced photosynthesis Toxic metabolites accumulate

Primary cause

Accelerated senescence and decay Increased water lossTissue discoloration Abnormal ripeningIncreased ethylene production Elevated respiration

SymptomsSymptoms

Stress-Induced Stress-Induced Ethylene Production is Ethylene Production is Another Response that Another Response that

Occurs Through a Occurs Through a Linear Sequence of Linear Sequence of

EventsEvents

ETHYLENE SYNTHESIS

Methionine

SAM

ACCACC

C2 H4

ACC Synthase

ACC Oxidase

MACC

Fruit ripeningFlower senescenceAuxin, WoundingChilling, DroughtFlooding

+

AVG, AOA

-Ripening, Ethylene

AnaerobiosisCobalt, > 35 °C

Free radical scavengers

+

-(CO2, O2 )

Hierarchical Response toabiotic stresses

Heat-shock Wounding Non-stressed

??DroughtAnaerobicSaltUV

Participants in this research

M.A. Mangrich F.A. Tomás-BarberánJ.G. Loaiza-Velarde

M.A. RitenourA. Rab

M. Cantwell G. Peiser

G. López-Gálvez

ABIOTIC STRESSES REDIRECT PROTEIN SYNTHESIS Mikal E. Saltveit Mann Laboratory, Department of...

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