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-- Introduction to Introduction to ‘‘OmicsOmics’’ and Systems Biology
Sixue Chen, Ph.D.
Department of Botany, Plant Molecular and Cellular Biology
Genetics Institute
E-mail: [email protected]
-- Demonstration of onDemonstration of on--line line ‘‘OmicsOmics’’ resourcesresources
-- Lab demonstration of modern ProteomicsLab demonstration of modern Proteomics
Introduction to Introduction to ‘‘OmicsOmics’’ and Systems Biology
Sixue Chen, Ph.D.
Department of Botany, Plant Molecular and Cellular Biology
Genetics Institute
E-mail: [email protected]
Large scale biology – ‘Omics’: revolution in screening important traits and creation of ‘in silico’ organisms
Chen lab research towards plant systems biology, an emerging discipline facilitates rational plant engineering
‘Omics’ modules – example of proteomics in addressing the differences in wheat (C3 plant) and corn (C4 plant)
OutlineOutline
What is modern and future biological sciences ?
Central Dogma of Molecular GeneticsCentral Dogma of Molecular Genetics
(The guiding principle that controls trait expression)
DNA(gene)
RNA
Protein Trait(or phenotype)
Transcription
Translation
Plant height
Seed shape
Traits and Phenotypes are Controlled by Molecular NetworksTraits and Phenotypes are Controlled by Molecular Networks
Trying to understand life without knowledge of biochemical network would be like trying to understand Shakespeare without knowledge of English grammar.
Our ProblemsOur Problems
• That one gene encodes one protein, which catalyzes one reaction and determines one phenotype is no longer the case.
• How to capture all molecules and their interactions, dynamics, regulations and turnover … ?
• How to determine the rate-limiting molecule and step ? How to predict ?
• Manipulating one gene can cause pleiotropic effects ?
.
.
.
Large scale biology – ‘Omics’: revolution in screening important traits and creation of ‘in silico’ organisms
Chen lab research towards plant systems biology, an emerging discipline facilitates rational plant engineering
‘Omics’ modules – example of proteomics in addressing the differences in wheat (C3 plant) and corn (C4 plant)
OutlineOutline
What is modern and future biological sciences ?
‘Omics’ and Systems Biology
• “Omics”– Genomics – the comprehensive
study of whole sets of genes & their interactions (DNA microarrays)
– Proteomics - the study of the full set of proteins encoded by a genome
– Metabolomics - the comprehensive study of the small molecules or metabolites
– Bioinformatics - the application of computer & statistical techniques to the management of biological information
Systems Biology-“An interdisciplinary approach for integrating experimental data with mathematical modeling tools to analyze & predict the behavior of biological systems.” (Henson, 2005)
Starvation: Importance of CassavaStarvation: Importance of Cassava
Cassava (Manihot esculenta) - yucca, manihot, tapioca• cultivated in tropics and sub-tropics for its edible storage root
• a major source of dietary energy for more than 700 million people
• source for a variety of food stuffs, animal feed and industrial products
• major component in micro-economies of more than 150 countries
Cassava tuberous roots developed from Cassava tuberous roots developed from fibrous rootsfibrous roots
• Tuberous roots develop form fibrous roots through massive cell division and differentiation of parenchyma cell of the secondary xylem
• Not all fibrous roots are designated for tuberous root formation
Proteomics is the approachProteomics is the approach
Protein map for comparison with other cultivars or tissue types
Compare physiological functions of the two types of roots
Identify targets for engineering
Fibrous Root
Tuberous Root
Protein Identification2D gel overlay
3D view of Fibrous Gel
3D view of Tuberous Gel
Protein Identification Using Liquid Chromatography Protein Identification Using Liquid Chromatography Mass SpectrometryMass Spectrometry
HPLC coupled online with 4000 QTRAP Mass SpectrometerHPLC coupled online with 4000 QTRAP Mass Spectrometer
Novel Targets for Biotechnological Application Novel Targets for Biotechnological Application
258257
258 257
263 263
Heat shock protein
BioCassava Plus project: it is proposed to increase the yield and protein content of cassava storage roots by four-fold
Galactose utilization in yeast
– Science 292: 929-934 (galactose utilization)Combines: literature knowledge, microarray, proteomics, visualization, and
network techniques to refine what is known about galactose utilization in yeast.
– Genome Res. 13: 244-253 (Genome scale network reconstruction)
Strategy
• For each gene or condition change (i.e. delete the gene) and measure the global effect on both mRNA and protein levels.
• Integrate mRNA and protein responses with the pathway model and with global network of protein interactions.
• Formulate new hypotheses to explain novel observations and refine models.
Galactose metabolism
- Science 292: 929-934
Expression measurements
10000
cDNAs
Hybridizationto microarray
mRNAextraction
Reverse transcriptionand fluorescent labeling of cDNA
WT control Mutant
Expression measurements
Microarray:
• a perturbed stain vs. wt + gal, 4 replicates
• statistics: maximum-likelihood estimation →997 significant genes → 16 clusters by self-organizing maps, each cluster contains genes with similar responses over all perturbations.
Expression measurements
Visualizing the data
gal4Δ
- Blue line (p-p); Yellow line (p-d); node diameter scales with the magnitude of cha
More Systems Biology to follow…
Systems Biology SchemeGenome(gene)
Transcriptome(mRNA)
Proteome(protein)
Metabolome(metabolite)
Phenotype
Representation of data in XML format- molecule identity and structure- quantification- functional status (modification and complex)- localization and dynamics
transcriptomics proteomics metabolomics
Statistical association, networking and modeling
New functions for genes, proteins and metabolites
In Silico Plant, Prediction and Rational Engineering
Simultation and phenotype explanation
Environment
Hypothesis modification
Large scale biology – ‘Omics’: revolution in screening important traits and creation of ‘in silico’ organisms
Chen lab research towards plant systems biology, an emerging discipline facilitates rational plant engineering
‘Omics’ modules – example of proteomics in addressing the differences in wheat (C3 plant) and corn (C4 plant)
OutlineOutline
What is modern and future biological sciences ?
Plants Produce Many Chemicals That Are Related to Our LifePlants Produce Many Chemicals That Are Related to Our Life
CH 2 CH
CH 2
S in ig r in A lly lg lu c o s in o la te
CH 2 CH
CH 2
CH 2
I. G lu c o s in o la te s d e rive d fro m m e th io n in e
G lu c o n a p in B u t-3 -e n ylg lu co s in o la te
CH 2 CH
CH 2
CH 2
CH 2
G lu c o b ra ss ic a n a p in P e n t-4 -e n ylg lu c o s in o la te
CH 2 CH
CH
CH 2O H
P ro g o itr in (2 R )-2 -H yd ro x yb u t-3 -e n ylg lu c o s in o la te
CH 2 CH
CH
CH 2O H
E p ip ro g o itr in (2 S )-2 -H yd ro xyb u t-3 -e n ylg lu co s in o la te
CH 2 CH
CH 2
CH
CH 2O H
N a p o le ife r in (2 R )-2 -H yd ro x yp e n t-4 -e n ylg lu c o s in o la te
CH 3 S CH 2
CH 2
CH 2
G lu co ib e rv ir in 3 -M e th ylth io p ro p ylg lu co s in o la te
CH 3 S CH 2
CH 2
CH 2
CH 2
G lu c o e ru c in 4 -M e th ylth io b u ty lg lu co s in o la te
CH 3 S CH 2
CH 2
CH 2
CH 2
CH 2
G lu co b e rte ro in 5 -M e th ylth io p e n ty lg lu co s in o la te
CH 3 S O CH 2
CH 2
CH 2
G lu co ib e r in 3 -M e th yls u lp h in y lp ro p ylg lu co s in o la te
CH 3 S O CH 2
CH 2
CH 2
CH 2
G lu c o ra p h a n in 4 -M e th yls u lp h in y lb u ty lg lu co s in o la te
CH 3 S O CH 2
CH 2
CH 2
CH 2
CH 2
G lu c o a lvs s in 5 -M e th ylsu lp h in ylp e n tylg lu c o s in o la te
CH 3 S O CH
CH
CH 2
CH 2
G lu co ra p h e n in 4 -M e th yls u lp h in y lb u t-3 -e n ylg lu c o s in o la teCH 3 S O 2 C
H 2
CH 2
CH 2
G lu c o c h e iro lin 3 -M e th ylsu lp h o n ylp ro p ylg lu c o s in o la te
G lu c o e ryso lin 4 -M e th ylsu lp h o n ylb u ty lg lu co s in o la te CH 3 S O 2 CH 2
CH 2
CH 2
CH 2
II. G lu c o s in o la te s d e rive d fro m p h e n y la la n in e
CH 2C
H 2
CH 2
G lu c o n a s tu rtiin P h e n e th ylg lu c o s in o la te
CH
CH 2O H
G lu c o b a rb a r in 2 -H yd ro x y-2 -p h e n yle th ylg lu c o s in o la teCH 2
OH
G lu c o le p ig ra m in m -H yd ro xyb e n zylg lu co s in o la te
CH 2
OHS in a lb in p -H yd ro x yb e n zylg lu c o s in o la teCH 2
C H 3 O
G lu c o lim n a n th in m -M e th o x yb e n zylg lu c o s in o la teCH 2
C H 3 OG lu c o a u b r ie tin p -M e th o x yb e n zylg lu co s in o la te
III. G lu c o s in o la te s d e rive d fro m tryp to p h a nG lu co b ra s s ic in In d o le -3 -ylm e th ylg lu co s in o la te
N
CH 2R 4
R 1
R 1 R 4 = H
N e o g lu co b ra s s ic in n -M e th o xyin d o le -3 -y lm e th ylg lu c o s in o la te R 1 = O C H 3 R 4 = H
S u lp h o g lu co b ra s s ic in n -S u lp h o in d o le -3 -y lm e th ylg lu c o s in o la te R 1 = S O 3- R 4 = H
4 -M e th o xyg lu c o b ra ss ic in 4 -M e th o x yin d o le -3 -ylm e th ylg lu co s in o la te R 4 = O C H 3
R 4 = O HR 1 = H
R 1 = H
-
O
N
S
O SO3
R
OH
OHOHOH
glucosinolate
FlavourPlant-insect
Defense
N/S nutritionN/S nutrition
Growth (IAA)Growth (IAA)
BiofumigationBiofumigation
“ I do not like broccoli. And I haven't liked it since I was a little kid and my mother made me eat it. And I'm President of the United States and I'm not going to eat any more broccoli.” – George Bush
Anticarcinogenicactivities
Goitrogenicproperty
Genetic Engineering Problem: Modulation of glucosinolatesaffects normal plant growth and development
35S::CYP83B1
ControlControlcyp79f1 Control35S::CYP79F2
cyp83a1
WT
atvamugt74WT
IG
A B C D
cyp79f1 cyp79f1
E HF
WT sur1
H
atvam3-4Wild type
thiohydroximate-O-sulfate
IAA
indole-3-acetonitrile
NIT
tryptophan
indole
indole-3-glycerolphosphate
indole glucosinolate
aldoxime
(n)homomethionine
methionine
S-alkylthiohydroximate
syringyllignin
sinapateesters
MAM
CYP79F1/F2
shikimatepathwaychorismateanthranilate
phenylalanine
cinnamic acid
lignin
flavonoids
coniferaldehyde/coniferyl alcohol
CCoAOMT
sinapaldehyde/sinapyl alcohol
COMT
SUR1
thiohydroximate
desulfoglucosinolate
aliphatic glucosinolate
aldoxime
S-alkylthiohydroximate
CYP79B2/B3
SUR1
thiohydroximate
desulfoglucosinolate
CYP83A1
S-GT S-GT
ST ST
dihydrocamalexic acid
CYP83B1
tryptamine
N-hydroxytryptamine
camalexin
CYP71B15
indole-3-pyruvic acid
indole-3-acetaldehyde
indole-3-acetaldehyde
TGG
AtDofMyb51 ATR1 IQD1 Myb28/29SLIM1
Sulfur deficiency
AP2
Wound, pathogen and insect
ET JA SA
Other factorse.g., K+
CYP71A13
- Yan and Chen (2008) Planta 226, 1343.
Proteomics: large scale analysis of proteins Proteomics: large scale analysis of proteins
Control Mutant
Protein expressionProtein expressionPhosphoproteomicsPhosphoproteomics
GlycoproteomicsGlycoproteomics
MetabolomicsMetabolomics: large: large--scale analysis of metabolitesscale analysis of metabolites
Statistical Analysis, Network and ModelingStatistical Analysis, Network and Modeling
Validation of targets and rational engineering.
““Golden RiceGolden Rice””Over 120 million children worldwide are deficient in vitamin A. Rice has been engineered to accumulate β-carotene, Incorporation of this trait into rice cultivars and widespread distribution could prevent 1 to 2 million
deaths each year.
Vitamin A deficiency is a serious problem• Causes blindness• Influences severity of diarrhea, measles
Large scale biology – ‘Omics’: revolution in screening important traits and creation of ‘in silico’ organisms
Chen lab research towards plant systems biology, an emerging discipline facilitates rational plant engineering
‘Omics’ modules – example of proteomics in addressing the
differences in wheat (C3 plant) and corn (C4 plant)
OutlineOutline
What is modern and future biological sciences ?