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Biochemistry ofCork Formation as a Stress Response
Cândido Pinto RicardoInês Chaves March 5th, 2009
Moore et al. 1998 Botany WCB/McGraw-Hill Companies
Secondary growth of dicot stems
Moore et al. 1998 Botany WCB/McGraw-Hill Companies
Secondary growth of dicot stems Axial section of cork oak tree
Silva et al 2005 International Materials Reviews vol. 6 pp 345
In the cork oak tree, the phellogen forms a continuous layer of cells, enveloping the tree trunk. Each year is produced a 2-3 mm thick layer of cork that adheres to that of the previous year.
A- Cork (Phellem)B- Subero-phellogenic changeC- PhellogeniumD- Liber tissueE- Liber wood changeF- Wood (Xylem)G- BarkH- Lenticular channelI- Area for stopper productionJ- Annual growth rings
www.winomagazine.com/blog2/?cat=8
CorkApplications
http://www.ccrc.uga.edu/~mao/intro/ouline.htm
Structure of plant cell wall
http://www.ccrc.uga.edu/~mao/intro/ouline.htm
T- Tertiary wallS- Secondary wallW- Extractables
(waxes, terpenes, sterols, etc)P- Primary wallM- Middle lamellaPo- Pore
Structure of cork oak cell wall
Silva et al 2005 International Materials Reviews vol. 6 pp 345
Structure of plant cell wall
T SuberinS
W P M
Po
Chemical composition (%):
Suberin 30-50Lignin 15-27Polysaccharides 12-25Extractables (waxes, terpenes, sterols, etc) 8-20Ash 2- 5Others 1- 5
Silva et al 2005 International Materials Reviews vol. 6:345
Cork from cork oak tree
SuberinComplex polymerTwo distinct domains:Polyphenolic and polyaliphatic
Lulai 2007 Skin-set, wound-healing and related effectsin Dick Vreugdenhi Ed. Potato Biology and Biotechnology Advances and Prespectives Elsevier, Amsterdam
Lignin structure
Phenolic precursors
Triose-PCalvinCycle
Pi
CO2
Erythrose-4-P
Sucrose
PEP
DAHP
Shikimate
Triose-P
PEP
Pi
Cytosol
Quinic acid
Quercitol
○
○
Glucose-6-P myo- Inositol-1-P
3-DehydroquinateNADH
3-Dehydroshikimate
Plastid
Triose-PCalvinCycle
Pi
CO2
Erythrose-4-P
Sucrose
PEP
DAHP
Phenylalanine
Shikimate
Triose-P
PEP
Pi
Chorismate
Cytosol
Quinic acid
Quercitol
○
○
Glucose-6-P myo- Inositol-1-P
3-DehydroquinateNADH
3-Dehydroshikimate
Arogenate
Tyrosine
Tryptophan
Plastid
Triose-PCalvinCycle
Pi
CO2
Erythrose-4-P
Sucrose
PEP
DAHP
Phenylalanine
Shikimate
p-Coumaric acid
Triose-P
PEP
Pi
Chorismate
Phenylalanine
FlavonoidsCytosol
Quinic acid
Quercitol
○
○
Glucose-6-P myo- Inositol-1-P
3-DehydroquinateNADH
3-Dehydroshikimate
Arogenate
Tyrosine
Tryptophan
CoumarinsStilbenes
p-Coumaryl alcoolCaffeic acid
Ferulic acid
Sinapic acid
Coniferyl alcool
Sinapyl alcool
Plastid
Triose-PCalvinCycle
Pi
CO2
Erythrose-4-P
Sucrose
PEP
DAHP
Phenylalanine
Shikimate
p-Coumaric acid
Triose-P
PEP
Pi
Chorismate
Phenylalanine
FlavonoidsCytosol
Quinic acid
Quercitol
○
○ LigninsLignans
Cork arom. domain
Glucose-6-P myo- Inositol-1-P
3-DehydroquinateNADH
3-Dehydroshikimate
Arogenate
Tyrosine
Tryptophan
CoumarinsStilbenes
p-Coumaryl alcoolCaffeic acid
Ferulic acid
Sinapic acid
Coniferyl alcool
Sinapyl alcool
Plastid
Triose-PCalvinCycle
Pi
CO2
Erythrose-4-P
Sucrose
PEP
DAHP
Phenylalanine
Shikimate
p-Coumaric acid
Triose-P
PEP
Pi
Chorismate
Phenylalanine
FlavonoidsCytosol
Quinic acid
Quercitol
○
○ LigninsLignans
Cork arom. domain
Glucose-6-P myo- Inositol-1-P
3-DehydroquinateNADH
3-Dehydroshikimate
Arogenate
Tyrosine
Tryptophan
CoumarinsStilbenes
p-Coumaryl alcoolCaffeic acid
Ferulic acid
Sinapic acid
Coniferyl alcool
Sinapyl alcool
● DQSDQS
DQS-Dehydroquinate synthase; CM-Chorismate mutase; PAL-Phenylalanine ammonia lyase; CAD-Cinnamoyl alcohol dehydrogenase;CinOR-Cinnamoyl oxiredutase; ChS-Chalcone synthase
● CMCM
●PALPAL
CADCAD●
CinORCinOR●
ChSChS●
Plastid
Aliphatic Precursors of Suberin
KAS- β-Ketoacyl-ACP synthetaseAT - Acyl tranferaseTE - Trans-enoylaseDS - DesaturasePC - Phosphatidyl-cholineTG -Triacilglicerol
Plastid
Endoplasmic reticulum
Plant Lipid Metabolism
1- Two dehydrogenase steps via ω-oxo acid interediate; 2- ω-Hydroxylation; 3- In-chain hydroxylation; 4- Epoxidation of the double bond, followed by hydration5- Fatty acid synthetase; 6- β-Ketoacyl-ACP synthetase II (KAS II) step of fatty acid synthetase; 7- Stearoyl-ACP Δ9-desaturase; 8- Fatty acid elongationACP-Acyl carrier protein
Harwood (1997) Plant Lipid Metabolism in Dey and Harborne Ed. Plant Biochemistry Academic Press, San Diego
Comparative analysis:phellem versus xylem
GenesProteins
Functional categories of genes contributing to cork formation
Soler et al. (2007) A Genomic Approach to Suberin Biosynthesis and Cork Differentiation. Plant Physiology 144:419–431
Differentially Expressed Proteins 3 IEF 10
MM
Carbohydratemetabolism
Energy
Secondarymetabolism Membrane transport
Stress/Defence
Unkown
Regulation/Signalling
10 ºC 28 ºC
15 day
30 day
Stress effects: Air Temperature (10 ºC and 28 ºC)
Triose-PCalvinCycle
Pi
CO2
Erythrose-4-P
Sucrose
PEP
DAHP
Phenylalanine
Shikimate
p-Coumaric acid
Triose-P
PEP
Pi
Chorismate
Phenylalanine
FlavonoidsCytosol
Quinic acid
Quercitol
○
○ LigninsLignans
Cork arom. domain
Glucose-6-P myo- Inositol-1-P
3-DehydroquinateNADH
3-Dehydroshikimate
Arogenate
Tyrosine
Tryptophan
CoumarinsStilbenes
p-Coumaryl alcoolCaffeic acid
Ferulic acid
Sinapic acid
Coniferyl alcool
Sinapyl alcool
● DQSDQS
DQS-Dehydroquinate synthase; CM-Chorismate mutase; PAL-Phenylalanine ammonia lyase; CAD-Cinnamoyl alcohol dehydrogenase;CinOR-Cinnamoyl oxiredutase; ChS-Chalcone synthase
● CMCM
●PALPAL
CADCAD●
CinORCinOR●
ChSChS●
Plastid
The suberized skin of potato tuberas a model to study cork metabolism
Structure of Potato Periderm
Lulai 2007 Skin-set, wound-healing and related effectsin Dick Vreugdenhi Ed. Potato Biology and Biotechnology Advances and Prespectives Elsevier, Amsterdam
Barel, G. et al. J. Exp. Bot. 2008 59:3347-3357
Potato tuber skin development
(A) Number of skin layers (suberized phellem cells) during tuber development
(B) Early stage in periderm development(C) Close-up of dividing phellogen cells(D) Mature skin following foliage removal Bar=200 µm
Cross-sections of tuber surface stained with Safranin O/Fast green and viewed by light (B–D, left panels) and UV (B–D, right panels) microscopy to examine tissue morphology and autofluorescence of suberized cells.
Barel, G. et al. J. Exp. Bot. 2008 59:3347-3357
Representative 2-DE images of skin and tuber storage parenchyma (flesh) at the developmental
stage of 8 weeks post-sprout-emergence
Barel, G. et al. J. Exp. Bot. 2008 59:3347-3357
Cell proliferation Oxidative stressActin (ACT) Ascorbate peroxidase 1 (APX1), cytosolicP23 tumor protein-like (P23/TCTP) Catalase isozyme 2 (CAT2)Proteasome {alpha}-7 subunit Catechol oxidase B, chloroplast precursorProteasome β-2A subunit Polyphenol oxidase (PPO)Translation init iation factor 5A-3Tubulin {alpha}-chain Plant defenceSignal transduction—cell wall Cysteine protease 1 (CYP1)Remorin (REM) Elicitor-inducible protein EIG-J7
Elicitor-inducible protein EIG-J7General metabolism Endochitinase 2 precursorUDP-glucose:protein transglucosylase (UPTG2) Endochitinase 2 precursorDisulphide-isomerase protein (PDI) PatatinTriosephosphate isomerase, (TPI) cytosolic isoform Patatin putative homologOxidative respiratory chain Patatin protein 07APFI (hypothetical protein F8G22.2) Pathogenesis-related protein 10 (PR-10)NADH-ubiquinone oxidoreductase 18 kDa subunit Pathogenesis-related protein 10 (PR-10)NADH:FMN oxidoreductase-like protein 2-Oxoglutarate-dependent dioxygenase (SPP2)
One-carbon (C1) metabolism Suberization/lignificationGlutamate-ammonia ligase (GS1) ACP-17 kDa β-hydroxyacyl-acyl carrier proteinSerine hydroxymethyltransferase 4 (SHMT4) Caffeoyl-CoA O-methyltransferase-5 (CCoAOMT-5)Methionine synthase (MS) Caffeoyl-CoA O-methyltransferase-6 (CCoAOMT-6)Abiotic and biotic stress Caffeoyl-CoA O-methyltransferase-3 (CCoAOMT-3)Plasma-membrane polypeptide (DREPP) Peroxidase (POD 18)
Peroxidase PER9-6 secretory (POD 20)Reference protein Peroxidase 136, class III , precursor (POD 9)Nascent polypeptide-associated complex NAC; UBA-like Peroxidase putative (POD 5)
Peroxidase, suberization-associated anionic peroxidase
List of proteins that accumulate differentially in potato tuber skin compared to tuber storage parenchyma
EVALUATION OF WOUND-HEALING PROCESSES IN POTATO TUBER TISSUE
Potato Slices
Day 0 Day 8Day 4
Potato Slices
Day 0 Day 8Day 4
Cellular dediferentiationGene expressionOxidative stress response
Suberin deposition
Wounding - Wound response - Healing
Periderm formation
Potato Slices
Day 0 Day 8Day 4
Cellular dediferentiationGene expressionOxidative stress response
Suberin deposition
Suberin detected
Wounding - Wound response - Healing
Periderm formation
Time-course of peroxidase and oxidase activity in wound-healing potato tubers
NADPH-dependent O2.– generation
Peroxidase activity
Razem et al. J.Exp. Botany, Vol. 54:935-941
Proteomics of the wound-healing process
3 10pI
Clustered mean expression profiles of differentially expressed proteins
Clustered mean expression profiles of differentially expressed proteins
Wound response
Clustered mean expression profiles of differentially expressed proteins
Periderm reconstruction
Clustered mean expression profiles of differentially expressed proteins
Periderm reinforcement
Cork FormationCellular Processes• Phellogen proliferation • Phellogen derived cells
– Phellem commitment– Cell expansion– Cell senescence– Suberin biosynthesis and waxes deposition– Cell death
Biochemical Processes• Cork results from 4 main secondary metabolic pathways:
– Acyl-lipids (aliphatic suberin domain)– Phenylpropanoids (cork aromatic components)– Isoprenoids (wax terpenes and sterols)– Flavonoids (tannins)
• Peroxidase activity fundamental (presence of H2O2)• Integration of biochemical process almost unkown