FOODCROPS.VN. 2016. Motoaki seki. RIKEN cassava initiative

1. World Congress on Root and Tuber Crops
January 18-22, 2016, Nanning, China
Plenary Session PS15, Jan. 22, 2016
RIKEN Cassava Initiative:
1. Collaboration with ASEAN countries and CIAT
2. Integrated Omic Analysis (Transcriptome,
Metabolome, Hormonome and Epigenome)
Motoaki Seki,Yoshio Takei (S09-05),Tetsuya Sakurai,
Tomoko Abe and Yoshinori Utsumi (SP06-22)
(RIKEN, Yokohama City Univ., JST CREST)

2. Cassava is an important crop in worldwide
-Usage and application of cassava biomass-
Foods & industrial materialUse as food and feed

3. Indonesia =
23.9 Mt/year
Thailand
= 22 Mt/year
China= 4.7 Mt/year
Brazil = 24.5 Mt/year
Colombia = 2.4 Mt/year
Paraguay= 2.6 Mt/year
Democratic Republic
of Congo = 15.0 Mt/year
Ghana = 13.5 Mt/year
Asia =
75.2 Mt/year
Africa =
121.0 Mt/year
South America
= 31.7 Mt/year
FAO STAT2010 (http://faostat.fao.org/)
Vietnam
= 8.5 Mt/year
Nigeria = 37.5 Mt/year
An Important Tropical Crop for Food Security, Poverty Reduction and
industrial material in many Asian and African countries
(an important source for a billion people’s food and income generation).
3
India =
10.0 Mt/year

4. π
Cassava production：265Mt (2007)→266Mt (2010)
Cassava production:58.2Mt (2004)→85.2Mt
(2010)
Cassava production：161Mt (2000)→239Mt (2010)
Japan
(RIKEN)
0.66Mt
（2014）
Ethanol industry
Cassava Relationship between ASEAN Counties and Japan
Citation
Agriculture & Livestock Industries Corporation(http://www.alic.go.jp/）
India
Thailand
(Mahidol Univ., DOA, KMUTT etc.;
PI: Dr. Naranganjavana)
Vietnam (AGI; PI: Dr. Ham)
Cambodia
Indonesia (LIPI, ILETRI)
China
Laos
Myanmar
Cambodia (Univ. of Battambang))
Cassava:

5. Mahidol Univ. (Thailand)
・Marker Breeding
(HB60XHN)
・CAD, CBB
CIAT (Colombia)
・Useful genetic
resources
・Molecular breeding
RIKEN (Japan)
・Identification of candidate genes using microarray
・Production of useful cassava by transformation
・Heavy-ion beam irradiation (with Dr. Abe, RIKEN
Nishina)
・Visit of Vietnamese Vice Prime Minister (May 22,
2013) and VAAS President (May 2, 2014) to RIKEN
・Selection of elite lines as pre-
commercial variety
AGI (Vietnam)
・Evaluation of newly
developed useful cassava
candidates in the field
Developing Contribution to
Green Innovation
Cassava International Collaborative Research
in e-ASIA program (Japan and ASEAN countries)

6. 1. Development of an integrative, functional-
genomics platform for cassava
2. Transcriptome Analysis (CAD etc.)
3. Heavy-Ion Beam Mutagenesis
4. Transformation
5. Integrated Omic Analysis (Transcriptome,
Metabolome and Hormonome) during tuberous
root development
6. Epigenetic Regulator (towards development of
stress-tolerant cassava)
Contents

7. ATG STOP
Genome DNA
mRNA
Partial cDNA
Full-length cDNA
Can’t produce
proteins
・Can produce
proteins
(Applicable to
transgenic plants)
Exon
Intron
Full-length cDNA is an useful tool
for basic and applied sciences.
・Useful for correct
genome annotation,
CRISPR design and
marker development.

8. Large-scale collection of Cassava full-length cDNAs
（Collaboration with CIAT)
・Treatment for isolation of full-length cDNA (abiotic stress) : drought、 heat、PPD、
heavy metal pollution（Al）
・EST: 20,000 full length cDNA (BMC Plant Biology 2007)
Treatment for isolation of full-length cDNA
Biotic stress : Mealybug, Whitefly, Mite, Bacterial blight and Root rots
Treatment for isolation of full-length cDNA
Biotic stress : Whitefly, Green mites, Mealybug, Hornworm, Bacterial blight
Other condition : Pesticide, fungicide, auxin, drought, fertilization water
2. MECU72(tolerance to whitefly)
1. MTAI16(elite cultivar in east-Asia）
3. MPER417-003（M. peruviana）（tolerance to whitefly and mealybug ）
4. Huay Bong 60（high yield cultivar）
5. Hanatee（low yield cultivar）
Meaybug treatment
CMC40 MPER417-003
・high yield
・high starch
・high cyanide ・tolerance to CAD
・low yield
・low starch
・low cyanide ・sensitive to CAD
Sakurai et al. (2007) BMC Plant Biol.
Fernando et al. (in preparation)

9. Identification of more than 30,000 genes from cassava.
Cultivars
Number of
clones
Number of tags
Number of mapped
tags
KU50
Sanger
19,968 35,400 34,432
MECU72 29,952 21,364 19,879
MPER417-003 19,968 15,626 15,309
MECU72
Roche/454
- 154,397 104,379
MPER417-003 - 471,934 397,932
MPER417-003
Illumina
- 51,919,340 12,387,186
Hanatee - 13,519,852 6,381,632
Huay Bong 60 - 107,583,892 39,497,396
Number of genes from Phytozome v10.1：30,666
Number of genes from full-length cDNA resources ：27,153
Number of novel genes ：more than 4,000
9
Sakurai et al. (2007) BMC Plant Biol.
Fernando et al. (in preparation)
Full-length sequences of about 7,000 KU50 FL-cDNAs
(with Dr. Iuchi, RIKEN BRC)
Improving Genome Annotation (with CIAT)
Isolation of about 27,000 cassava Full-length cDNAs.

10. Development of Cassava Oligoarray
(containing about 30,000 genes)
2010~: 30,000 gene
Agilent oligoarray
developed
September: >15,000 gene
Agilent oligoarray
developed
Agilent 20K oligoarray
experiments
(in progress)
Agilent 30K
oligoarray experiments
were done
2009 2010 2011
2nd custom 30K oligoarray
Futures:
•>30,000 gene proves on a chip
• More biotic-related genes
• Genes from wild species
10
Utsumi et al. (2012) DNA Res.
Sojikul et al. (2015) Plant Mol. Biol.
Utsumi et al. (revised)

11. Construction of cassava database based on FL-cDNA information
(Cassava Online Archive, http://cassava.psc.riken.jp/)
（Sakurai et al. 2013
PLoS One）
11
Integration with
outer database
Collection of
FL-cDNA Seq
information
Results
BLAST
Top page
Keyword
Genome
Browser

12. 1. Development of an integrative, functional-
genomics platform for cassava
2. Transcriptome Analysis (CAD etc.)
3. Heavy-Ion Beam Mutagenesis
4. Transformation
5. Integrated Omic Analysis (Transcriptome,
Metabolome and Hormonome) during tuberous
root development
6. Epigenetic Regulator (towards development of
stress-tolerant cassava)
Contents

13. Huay Bong 60 (HB60) Hanatee (HN)
・High yield
2006
・Resistant to
Cassava anthracnose disease (CAD)
・High starch content
・High HCN content
・Low yield
・Sensitive to CAD
・Low starch content
・Low HCN content
Cassava Marker Breeding in Thailand
-Development of F1 population (HB60 X HN)-
(Collaboration with Mahidol Univ.)

14. Linkage map analysis : Results
Fresh root yield
Starch content
Cyanogen content
Starch properties
CAD resistance
Marker Breeding (Collaboration with Mahidol Univ.)
14

15. ・Deep sequencing revealed 595 SNPs in genomic regions including candidate
genes involved in CAD resistance and starch content. (in collaboration with
Mahidol Univ.)
・Identification of 10,546 SNPs in 3,252 genes
（123 SNPs on suar and starch metabolism pathway genes）

17. 17
Identification of candidate genes of CAD tolerance by
microarray analysis (Collaboration with Mahidol Univ.)
A
Kunkeaw et al. (2010) Australas Plant Pathol, 39:547-550
B
But resistance mechanism for CAD is still unknown…
HB60 KU50HN
HNKU50 HB60

18. Venn diagram analysis of the genes with higher (A) and
lower (B) expression in HB60 and HN.
18
Identification of the genes with higher
expression in a CAD-tolerant cultivar (HB60) .
Utsumi et al. revised)

19. GO term of the genes with higher
expression in HB60 compared with
HN under non-treatment condition
Various immune systems
are involved in CAD
resistance in HB60.
GO term of the genes with higher
expression in HB60 compared with
HN at 72 h after CAD infection
Utsumi et al. revised

20. 1. Development of an integrative, functional-
genomics platform for cassava
2. Transcriptome Analysis (CAD etc.)
3. Heavy-Ion Beam Mutagenesis
4. Transformation
5. Integrated Omic Analysis (Transcriptome,
Metabolome and Hormonome) during tuberous
root development
6. Epigenetic Regulator (towards development of
stress-tolerant cassava)
Contents

21. Cassava Breeding by Heavy-Ion Irradiation (non-GM)
(Collaboration with AGI, RIKEN Nishina-Center )
Cassava Fruit
Heavy-ion
irradiation
(50〜150 Gy)
Screening
Collect the
embryo
Germination
on the media
21
Collect the seeds

22. About 1,000 heavy ion beam-irradiated
cassava KU50 plants have been grown in AGI (Vietnam)

23. Pandurate
leaflobe Bloom in Hanoi
5 leaf lobes 9 leaf lobes
Red petiole Green petiole
Purple apical leaf (parent) Dark green apical leaf Light green apical leafHigh starch (30%)
Phenotype variations in about 1,000 ion beam-
irradiated lines

24. Can analyze
more than
30,000 gene
expression
(3) Preparation of
custom oligomicroarray
(1) Isolation and sequencing
of about 27,000 Full-Length
cDNAs
(2) Cassava
database
(4) Marker Breeding
(6) Transgenic Cassava
(5) Heavy-Ion
Irradiation Breeding
Development of Cassava Research Platform
towards Molecular Breeding

25. 1. Development of an integrative, functional-
genomics platform for cassava
2. Transcriptome Analysis (CAD etc.)
3. Heavy-Ion Beam Mutagenesis
4. Transformation
5. Integrated Omic Analysis (Transcriptome,
Metabolome and Hormonome) during tuberous
root development
6. Epigenetic Regulator (towards development of
stress-tolerant cassava)
Contents

26. Friable embryogenic callus(FEC)
Transgenic Plantlets
Agrobacterium infection
In Vitro mother plantlets
Axillary buds or small leaves
Somatic embryogenic callus
Transformed FEC
Cotyledon
fragments
Transgenic shoot
initials
Agrobacterium infection
Agrobacterium-mediated cassava transformation
of friable embryogenic calli (FEC)
PCR and GFP
selection
Control leaf
Regeneration from FEC
Somatic embryogenic callus

27. List of media for FEC production in cassava
Genotype
Successful genotype to FEC
prodaction
Media for FEC production
Sucrose
concentration
Auxin Reference
1. TMS60444 and M.COL. 1505 TMS60444 and M.COL. 1505
MS, ½MS, ,MS(-NH4), MS(-NH4NO3), SH,
N6, NN, GD, WPM and B6
2% 12 mg/l picloram Taylor et al. 1996 (Nat. Biotechnol)
2. TMS60444 TMS60444 GD 2% 10 mg/l picloram Raemakers et al. 1996 (Mol. Breeding)
3. TMS60444 and MCOL22 TMS60444 and MCOL22 GD 2% 12 mg/l picloram Zhang et al. 2000 (Ph.D. thesis in ETH)
4. TMS60444, Adira 4, R60, R90, Thai5, M7,
Mcol22, Adira 1, L11 and Gading
TMS60444, Adira 4, R60, R90,
Thai5*1
and M7*1 GD 6%
6 mg/l picloram
6 mg/l NAA
Raemakers et al. 2001 (Euphytica)
5. TMS60444 TMS60444 GD 2% 12 mg/l picloram Schreuder et al. 2001 (Euphytica)
6. Bujá Preta, Rosinha Bujá Preta, Rosinha GD 2% 12 mg/l picloram Ibrahim et al. 2008 (Afri J Biotechnol)
7. TMS60444 TMS60444 GD 2% 12 mg/l picloram Bull et al. 2009 (Nat Protoc.)
8. T200, AR9-18, MTAI16, CR25-4, CM523-7,
BRA1183, MCOL2261 and SM707-17
- MS*2
2% 12 mg/l picloram Rossin et al. 2010 (S. Afr. J. Bot.)
9. TME 3, TME 7 and TME 14 TME 3, TME 7 and TME 14 GD 2% 12 mg/l picloram Zainuddin et al. 2012 (Plant Methods)
10. TMS60444 and T200 TMS60444 and T200 GD 2% 12 mg/l picloram Chetty et al. 2013 (N Biotechnol.)
11. Ebwanatereka, Serere and Kibandameno
Ebwanatereka, Serere and
Kibandameno
GD with different concentrations of tyrosine
(125, 250 and 500 µM)
2% 50 µM picloram Nyaboga et al. 2013 (Frontiers in plant sci.)
12. Aladu, Ebwanateraka, 60444 Aladu, Ebwanateraka, 60444 GD with 500 µM tyrosine 2% 50 µM picloram Apio et al. 2015 (African J Biotechnol)
13. TME14 TME14 GD 2% 12 mg/l picloram Nyaboga et al. 2015 (Frontiers in plant sci.)
*1 Thai5 and M7 were classified as relatively difficult lines to produce FEC. The efficiency of FEC production is about 1%.
*2 The production of somatic embryo from all genotypes was reported in this manuscript.

28. Media
TMS60444 KU50
Number of somatic
embryo
Number of colony
of friable
embryogenic callus
Percentage
(%)
Number of
organized callus
Number of colony
of friable
embryogenic
callus after
cultured during 4
weeks
Percentage
(%)
Murashige and Skoog medium (MS) 13 4 31 27 0 0
MS (×1/100 NO3) medium 12 0 0 25 0 0
MS (×1/100 NH4) medium 12 0 0 25 0 0
Media X exp1 15 15 100 24 0 0
exp2 70 51 73 - - -
exp3 56 29 52 - - -
MS (-Zn) medium 13 1 8 30 0 0
MS (-Br) medium 13 1 8 21 0 0
Gresshoff & Doy basal medium exp1 29 9 31 50 0 0
exp2 60 22 37 - - -
exp3 59 20 34 - - -
McCow'n woody plant medium 16 6 38 25 0 0
Chu N6 medium 14 4 29 28 0 0
Gamborg B5 medium 15 5 33 25 0 0
Optimization of media for FEC production in cassava
Ha, Utsumi et al. in preparation

29. Workflow of cassava transformation
Ha, Utsumi et al. in preparation

30. Preparation of in vitro plantlets
from Vietnamese local cultivars
Multiplication of in vitro cassava plant
Candidate cassava cultivars
1 KM140 KM98-1 x KM36
2 KM325 SC5 x SC5
3 NA1 MIF
4 KM397 KM108-9-1 x KM219
5 HL2004-28 (GM444-2 x GM444-2) x XVP
6 TaXanhDB Unknown
7 Rayong11 R5 x OMR29-20-118
8 KM987 SM1717 x CM321-188 [AGI, VNM]
9 TCĐB SC5 x SC5 [AGI, VNM]
10 TĐĐB Unknown [AGI, VNM]
11 KU50 Rayong1 x Rayong90
12 TMS60444 Unknown [IITA, NGA]

31. Selection and screening of Asian cultivars
based on efficiency of FEC production
KU50 callus
on-going step…

32. Qualitative and Quantitative Improvement of Cassava Biomass using
Agrobacterium-mediated Cassava Transformation System
8 constructs
→Food security and stable
biomass supply
→Increase of non-food biomass →Industrial material
Quantity Quality
9 constructs
Improvement of photosynthesis
Increased tuberous roots
Early bulking
Early flowering
Stress tolerance
32

33. What is Fructose 1,6-bisphosphate Aldolase (FBA)?
①Fructose 1,6-bisphosphate ⇄ Dihydroxyacetone phosphate ＋Glyceraldehyde 3- phosphate
②Sedoheptulose 1,7-bisphosphate ⇄ Dihydroxyacetone phosphate + erythrose 4-phosphate
 Viewpoint
‐ Bypass of the pathway
between Calvin Cycle and
Sucrose transport.
‐ One of the key enzyme of
photosynthesis
‐ AtFBA3 is localized in
Plastid and Cytoplasm
The Plant Journal(2010) 61,1067-1091
Mark Stitt et al.
Arabidopsis and Primary photosynthetic metabolism
– more than the icing on the cake.
①
② ①
Reaction

34. Aldolase is a candidate gene for improving the
photosynthetic carbon fixation
Photosynthesis Research 75: 1-10
2003
Christine A. Raines
The Calvin cycle revisited
Aldolase is candidate of engineering to improve photosynthetic carbon
fixation.

35. Aldolaseactivity(μunit/μg/min)
AtFBA3-ox cassava plants has 2.0-
5.9-fold higher aldolase activities
than WT.
WT AtFBA3-
ox C8
AtFBA3-
ox E6
AtFBA3-
ox E3
×103Copynumber/ngtotalRNA
Gene expression of AtFBA3 and
Aldolase Activity in Leaves
AtFBA3-ox cassava plants has
higher gene expression than WT.
Sample is grown in Green house (sun light, 28℃) and collected from 2nd to 4th leaves from top of the stem at 10:00-12:00 .
Mean and error bar indicate the average value and standard deviation with each triplicate experiments using three biological replicates
WT AtFBA3-
ox C8
AtFBA3-
ox E6
AtFBA3-
ox E3
Gene expression Aldolase activity
(Takei et al. in prep.)

36. Red : p<0.05 Blue : p<0.1 by T-test
The tuber yield of FBA OX lines is increased
under 400 ppm CO2 condition.
(Takei et al. in prep.)

37. ＲR
RIKEN CSRS has established integrated Omic Analysis Platform.
Molecular Analysis to elucidate the mechanism is in progress.
Metabolome
Hormonome
Transcriptome and
Epigenome
Agilent Microarray Scanner
Ion Proton
SOLiD5500

38. 1. Development of an integrative, functional-
genomics platform for cassava
2. Transcriptome Analysis (CAD etc.)
3. Heavy-Ion Beam Mutagenesis
4. Transformation
5. Integrated Omic Analysis (Transcriptome,
Metabolome and Hormonome) during tuberous
root development
6. Epigenetic Regulator (towards development of
stress-tolerant cassava)
Contents

39. 39
Table 1. The sampling plan during developing stage.
4 weeks 6 weeks 8 weeks 10 weeks 12 weeks
1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 Total
Fibrous* Parenchyma
○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ 15
Cortex
Intermediate
Parenchyma
- - - - - -
○ ○ ○ ○ ○ ○ ○ ○ ○ 9
Cortex ○ ○ ○ ○ ○ ○ ○ ○ ○ 9
Storage
Parenchyma
- - - - - -
○ ○ ○ ○ ○ ○ ○ ○ ○ 9
Cortex ○ ○ ○ ○ ○ ○ ○ ○ ○ 9
Top of leaves ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ ○ 15
Total 66
: Metabolome, Transcriptome : Hormonome
Fibrous root； Diameter <2mm
Intermediate root；Diameter 2-5mm
Storage root; Diameter > 5mm
Cortex
Parenchyma
Integrated Omic Analysis (Transcriptome, Metabolome and
Hormonome) during Tuberous Root Development (with Mahidol Univ.)
(unpublished, Utsumi et al.)
39

40. Starch content in root tuber samples
Starch biosynthesis has started already in fibrous root from F8 and F12
Starchcontent
(mg/mgF.W.)
(Utsumi et al. in prep.)

41. Unknown
signal
ABA ↑
GA −
IAA −
IAAsp −
iP ↑
tZ ↑
cZ ↑
Sucrose↑
G6P↑
F6P↑
AMP−
ABA ↓
GA ↑
IAA −
IAAsp ↓
iP ↑
tZ ↑
cZ ↑
Sucrose↑
G6P↑
F6P↑
AMP↑
Sucrose↑
G6P↑
F6P↑
AMP−
Parencyma(0.02%) (3-5%)
(10-18%)
1. Post-embryonic
development
2. Response to
stimulus
3. Phosphorylation
4. Response to
chemical stimulus
5. Reproductive
process in a
multicellular
organism
1. Response to
stimulus
2. Post-embryonic
development
3. Glucan
biosynthetic
process
4. Cellular glucan
metabolic
process
5. Glucan metabolic
process
Metabolites
Plant
hormones
Transcriptome
Root state
(Starch content)
Fibrous StoragePre-storage
Integrated Omic Analysis during Tuberous Root
Development in Cassava (Utsumi et al. in prep.)
Intermediate
Stimulation
to plant
hormone
and sugar Parencyma
(8-15%)
ABA↓
GA −
IAA −
IAAsp ↓
iP ↑
tZ ↑
cZ ↓
Cortex(5-9%) Cortex(5-9%)
ABA ↓
GA −
IAA −
IAAsp ↓
iP ↑
tZ ↑
cZ ↓
Sucrose↑
G6P↑
F6P↑
AMP↑
Sucrose↑
G6P↑
F6P↑
AMP−
(unpublished, Utsumi and Seki et al.)
41
1. Post-embryonic
development
2. Photosynthesis
3. Cellular glucan
metabolic
process
4. Glucan metabolic
process
5. Response to
stimulus
ABA ↓
GA ↑
IAA −
IAAsp ↓
iP ↑
tZ ↑
cZ ↑
1. Response to
stimulus
2. Post-embryonic
development
3. Glucan
biosynthetic
process
4. Cellular glucan
metabolic
process
5. Glucan metabolic
process
1. Post-embryonic
development
2. Photosynthesis
3. Cellular glucan
metabolic
process
4. Glucan
metabolic
process
5. Response to
stimulus

42. G1P
L-kestose
Sucrose
F6P
2-1,3-aminopiperidin
l-arginine
l-ornitine
A
B
C
D
F
E
G
HI
J
K
L
G6P
l-serine
4-aminobutyric acid
beta-alanine
Maltotriose
UMP
AMP
l-aspartic acid
l-glutamic acid
Glycolic acid
CMP
trans-zeatin
Heatmap of 154 metabolites
increasing tuberization process
Oxalic acid
l-threonic acid
l-cysteine
Stigmasterol
5-aminolevulinic acid
3-ureidopropionic acid
Threonic acid-1,4-lactone
d-gluconic acid
F8
I8_C
I8_P
M8_C
M8_P
F12
I12_C
I12_P
M12_C
M12_P
d-maltose
Metabolisms involved in sugars and nucleotide increased in all
tissues in comparison to fibrous roots at 4 weeks after cutting.
(Utsumi et al. in prep.)

43. 1. Development of an integrative, functional-
genomics platform for cassava
2. Transcriptome Analysis (CAD etc.)
3. Heavy-Ion Beam Mutagenesis
4. Transformation
5. Integrated Omic Analysis (Transcriptome,
Metabolome and Hormonome) during tuberous
root development
6. Epigenetic Regulator (towards development of
stress-tolerant cassava)
Contents

44. 44
Area with high-salinity stress-affected damage （Sparks 1995）
High-salinity stress problems have been
reported in North East of Thailand and
Indonesia.
Area with drought stress-affected damage （Tottori Univ., Arid Region Research Center)
Areas with High-salinity or drought stress-affected damage occur in the world.
Asia =
75.2 Mt/year
Africa =
121.0 Mt/year
South America
= 31.7 Mt/year
Indonesia =
23.9 Mt/year
Thailand
= 22 Mt/year
China= 4.7 Mt/year
Brazil = 24.5 Mt/year
Colombia = 2.4 Mt/year
Paraguay= 2.6 Mt/year Democratic Republic
of Congo = 15.0 Mt/year
Ghana
= 13.5 Mt/year
FAO STAT2010 (http://faostat.fao.org/)
Vietnam
= 8.5 Mt/year
Nigeria = 37.5 Mt/year
India =
10.0 Mt/year
Cassava Production
We need to develop the cassava plants tolerant to the abiotic stresses!

45. 45
Salinity-accumulating Soil : 28,970 km2
Area with salt damage : 2,830 km2
鉱床岩塩 : 約 18 兆t
鉱床カリウム : 約 25 億t
岩塩層厚 : 約 100 m 〜 150 m
岩塩層深度 : 地下約 150 m 〜 300 m
(参考)
地盤工学会「土と基礎」,2007.3
PIPATPONGSA THIRAPONG (東京工業大学助教授)
飯 塚 敦 (神戸大学教授)
河 井 克 之 (神戸大学助手)
Area with high-salinity stress-affected damage in Thailand
North-east
of Thailand
Nakhon Ratchasima
Sakon nakhon
Bangkok

46. 46
(http://www.ne.jp/asahi/vietnam/agriculture/environment/region.htm#31)
World Bank Report (Dasgupta et al. 2007):Vietnam is
one of the countries that are affected most
seriously by climate changes、海面が 1 m上昇すると、
人口のおよそ 10%が影響を 受けるとされている。ベトナ
ムの天然資源環境省は、2009 年、「ベトナムにおける気
候変動と海面上昇に関するシナリオ」(Bo Tai Nguyen va
Moi Truong 2009)を作成し、各省がこのシナリオにもと
づき、堤防の建設など、具体的な対策を講じはじめてい
る。2012 年に最新のシナリオが公表されたが、これによ
ると、気候変動の影響を 3 段階に分け、影響力が「中」
であった場合、It is predicted that if the sea
level will rise 1 m, Mekong Delta area will
lose 39% of the Area and if the sea level
will rise 2 m,more than 92% of the area will
be lost (Nguoi Lao Dong 2012).
Drought stress problem occurs.
Areas with drought- and/or high-salinity stress-
affected damage (might occur in the future) in Vietnam
Similar situation occurs in India.

47. Plant Cell
Signal Perception and
Signal Transduction
ABA
Stress ResponseSalt
ABA
independent
Regulatory Protein
Functional Protein
Non-coding RNAs
RNA regulation
Epigenetic regulation
and Tolerance
Drought Cold
Heat
Drought Cold
High
Heatsalt
Small peptides

48. +
+
-- Repress
Alteration of histone modifications
Active-mark
Repressive-mark
HAT
HDAC
HMT
H3K27m3
H3K9ac
H3K4m3
HAT: Histone acetyl transferase
HDAC: Histone deacetylase
HMT: Histone methyl transferase
Transcription
Relaxing
Packing
HDM: Histone demethylase
HDM
HMT
Histone modification enzymes
Alteration of histone modifications is
deeply and universally correlated with the gene regulations in eukaryotes.

49. Acetylation
HDAC
deacetylation
Ac
AcAc
AcAc
Ac
AcAc
AcAc
Ac
Ac AcAc
Ac
Activation of the genes
involved in environmental stress tolerance
遺伝子発現
OFF
Gene Expression
ON
HDAC
deacetylation
HDAC
inhibitor
Histone Acetylation is involved
in activation of gene expression.

50. Ky-2
Ac Ac
HDAC
Ky-2, an HDAC inhibitor, enhances high-salinity
stress tolerance in Arabidopsis.
×
SOS1
Transcription ON
SOS1
Na+ Na+
SOS1
Na+
SOS1
SOS1Na+
Na+
Na+
Na+
Sako et al. (2016) Plant Cell Physiol.

51. 4℃ 22℃ (16h-L/8h-D)
9 day after
germination (DAG)
0 4 5
5 HDAC inhibitors enhancing high-salinity stress tolerance
0
10
20
30
40
50
60
70
80
90
100
DMSO JNJ MC1568 MS275 SAHA TSA
Survival rate (%)
(n=15, 3 biological replicates)
LBH5891 2 3 4 5
Ueda et al. (in preparation)

52. HDAC inhibitor 1-
treated plants + NaCl HDAC inhibitor 1-
treated plants + NaCl
HDAC inhibitor
1-treated plants
Control
HDAC inhibitor
1-treated plants
Control
Shoot (mg)Shoot (mg)
Root(mg)
Root(mg)
HDAC inhibitor 1 enhances high-salinity stress tolerance in cassava.

53. Transcriptome analysis identified many candidate genes of
HDAC inhibitor-mediated high-salinity stress tolerance
Genes with higher expression
in roots of HDAC Inhibitor 1-
treated plants (not high-salinity
stress-inducible)
Genes with lower expression
in roots of HDAC Inhibitor 1-
treated plants (high-salinity
stress-inducible)
NoInhibitor(Control)
NoInhibitor+2hNaCl
NoInhibitor+24hNaCl
24hInhibitor
24hInhibitor+2hNaCl
24hInhibitor+24hNaCl

54. ＲR
RIKEN CSRS has established integrated Omic Analysis Platform.
Molecular Analysis to elucidate the mechanism is in progress.
Metabolome
Hormonome
Transcriptome and
Epigenome
Agilent Microarray Scanner
Ion Proton
SOLiD5500

55. Japan
Kyushu Univ.
(PI: Dr. Takasu)
RIKEN
Tokyo Univ. Agr.
Univ. Tokyo
Nagoya Univ.
Vietnam
AGI
Main Res. Inst.
Sub Res. Inst.
Tested Cassava Field
Rayong Field Crop Res. Cent.
Thailand
Hung Loc Agr. Cent.
Univ. of Battambang
(UBB), Cambodia
55
JST-JICA SATREPS project (2016-2021) :
Development and dissemination of sustainable production system based on
invasive pest management of cassava in Vietnam, Cambodia and Thailand

56. 5656
Ajinomoto Co.
Useful biomass,
such as amino
acids
Fertilizers,
Nutrient
materials
Wastes
（Rich in nutrients,
such as nitrogen）
・Identification of
useful genes.
・Development of useful
cassava by transformation,
chemicals and heavy ion
beam irradiation..
Develop to field
Use of stems for propagation
Cassava starch
Cassava pulp
Cellulose Film
Spray on
the crops
Harvest of tuber roots
Foods
Sweetener
Industrial raw materials
Materials for food processing
Fermentation
Decompose into glucose
Environmentally friendly
reutilization
Green Innovation using Cassava
・Cooperation with Japanese Companies
・Sustainable Human Life through Stable
Supply of Sugars with Low Price
for Foods and Biomaterials

57. Collaborators
RIKEN CSRS
Yoshinori Utsumi (SP06-22)
Chikako Utsumi
Yoshio Takei (S09-05)
Vu The Ha
Onsaya Patanun
Minoru Ueda
Kaori Sako
Minoru Yoshida
Hitoshi Sakakibara
Kazuki Saito
Atsushi Fukushima
Miyako Kusano
Akihiro Matsui
Maho Tanaka
Junko Ishida
Jong-Myong Kim
Tetsuya Sakurai
Satoshi Iuchi
Masatomo Kobayashi
Ken Shirasu
Kazuo Shinozaki
Thailand, Mahidol University
Jarunya Narangajavana
Kanokporn Triwitayakorn
Punchapat Sojikul
Supajit Sraphet
Sukhuman Whankew
DOA
Opas Boonseng
Amonrat Kitjaideaw
King Mongkut University of
Technology at Thonburi
Treenut Saithong
Saowalak Kalapanulak
Research Institute for Biological
Sciences, Okayama (RIBS)
Ken'ichi Ogawa
Yoshihiro Narusaka
Mari Narusaka
CIAT
Manabu Ishitani
Joe Tohme
Yokohama City University
Hiroyuki Tsuji
Nara Institute of Science and
Technology (NAIST)
Ko Shimamoto
Masaaki Umeda
RIKEN Nishina Center
Tomoko Abe
Tomonari Hirano
Vietnam, AGI
Ham Huy Le
Dong Van Nguyen
Vu Anh Nguyen

58. Thank you for your attention
58