Workshop Argentina-Japan “Bioscience and Biotechnology for the Promotion of Agriculture and Food Production” -August 3rd to 7th 2009- Hisanori Kato Endowed Chair of “Food for Life” Organization for Interdisciplinary Research Projects The University of Tokyo 1
Japan 日本 Life Expectancy at Birth (2008, by the Japanese Ministry of Health, Labor, and Welfare) Female: 86.05 years (the longest in the world) Male: 79.29 years (the 4 th in the world) 2
Japanese Blessed with a Variety of Healthy Traditional Foods Abundant Experience on Functional Food Factors Accumulated research data and experience will help health promotion of other countries!! 3
Functional Food in Japan – Food for Specified Health Uses (FOSHU) Nature , 364, 180 (1993)
Omics Analyses in Food and Nutrition Science Transcriptomics Metabolomics Spec #1[BP = 829.7, 24053] 829.7285 2.4E 100 90 Proteomics 80 881.7770 70 1378.3245 2191.8588 % Intensity 60 50 2222.0081 1799.7345 40 785.6865 1014.0126 1060.6566 30 833.5817 2279.1555 20 1927.8366 2145.8169 1088.0897 1696.5836 871.5511 2198.8981 1051.0375 2483.3095 2869.5020 1679.5564 1881.8104 10 709.6359 1412.3932 2175.0235 0 0 700 1260 1820 2380 2940 3500 Mass (m/z)
Effects of Dietary Protein Deprivation and Amino Acid Deprivation on mRNA Levels of IGFBP-1 Gene GROWTH RETARDATION 5 Rat Liver Hepatoma Cells 4 Relative mRNA Levels Relative mRNA Levels 6 5 3 4 2 3 2 1 1 0 0 AA+ AA- 12C PF （ 12 % Casein Diet ）（ Protein-free Diet ） Complete Amino Acid- Medium Deprived (12 h) 6
Transcriptome Analysis of the Effect of Food Food ・ Intake of Nutrients Control ・ Functional Food Factors Animals ・ Excessive Intake Comprehensive Evaluation of the Function and Safety Cells Exhaustive Analysis of Gene RNA Extraction Expression Treated x10 x5 x1/10 Control DNA Microarray Analysis x2 x1/5 x1/2
The World of OMICS Target Omics Discipline Nutritional Omics Nutrigenomics Genes Genomics (narrow sense) Transcripts Transcriptomics Nutritranscriptomics Proteins Proteomics Nutriproteomics Metabolites Metabolomics Nutrimetabolomics · · · · · · · · · · · · · · · Nutrigenomics Nutriomics 8
Other OMICS Epigenomics (Chromatin Modification) Interactomics (Protein-Protein Interaction) Toponomics (Cellular Localization) (Localizomics) Fluxomics (Cellular Flux of Metabolites) Lipidomics (Lipids) Glycomics (Sugars) CAUTION: Economics and Comics Do NOT · · · Belong to the Omics Family
Use of DNA Microarray 1.Nutritional Intervention 2.Known Function of a Food Factor 3. A Food Material or Component of Interest DNA Microarray and Other Omics Analyses Understanding Whole Response Understanding Mechanisms Unknown Functionality 10
12% Casein 12% Gluten Protein Free （ 12C) （ 12G) （ PF) 140 (+Met 2) 0 0 Casein 0 150 0 Gluten 656 648 798 Cornstarch 50 50 50 Mineral Mix 40 40 40 Vitamin Mix 10 10 10 Cellurose 100 100 100 2 2 2 Choline chloride Essential Amino Acid Pattern of Gluten Rat Body Weight Ile 240 12C Leu 220 12G Lys PF 200 Met+Cys 180 Phe+Tyr 160 c Thr 140 Trp 120 -4 -2 0 2 4 6 8 Val 11 0 50 100 150%
Object To obtain a comprehensive view of the changes in gene expression in response to the alteration of protein nutrition Method Male Wistar rats ( 5 wk old) 12% Casein RNA extraction 12% Casein Protein Free (Liver, Muscle,,,) 12% Gluten - 6 0 7 (Day) Rat U34A Genome Feeding (10:00~18:00) Array (Affymetrix) 12
Ingestion of Gluten Up- -regulates Genes for regulates Genes for Ingestion of Gluten Up Both Synthesis and Catabolism of Cholesterol Both Synthesis and Catabolism of Cholesterol 2 Acetyl-CoA Lanosterol Lanosterol 14-demethylase (3.2) Acetoacetyl-CoA 14-demethyl lanosterol HMG-CoA synthase (5.4) HMG-CoA Dehydrocholesterol HMG-CoA reductase (4.4) Dehydrocholesterol reductase Mevalonate (3.2) Squalene synthetase Cholesterol (4.4) Cholesterol Squalene 7- α hydroxylase Squalene eposidase (2.5) (5.8) 7 α -Hydroxycholesterol Squalene epoxide
Protein Nutrition and Skin Gene Expression Protein Nutrition and Skin Gene Expression <day> -4 -3 -2 0 1 7 8 12% Casein Diet 12C 12% 12% 12% Gluten + Lys +Thr (12GLT) 12GLT Pellet Casein Casein Diet 12% Gluten (12G) 12G Diet Diet Protein- Free Diet (PF) PF 1.5 h 10:00 ～ 18:00 24 h Body Weight Excision of Skin DNA Microarray (n=4) Day
Professional Program for Agricultural Bioinformatics (Tokyo Univ.) 15
Choice of Normalization Method Choice of Normalization Method DFW MAS Body Weight RMA qFARMS
Other Ongoing Projects (transcriptomics) Deficiency and Excess of Amino Acids Specific Proteins and Pepetides Protein Intake Levels at Various Life Stage Effect of Dietary Factor on Longevity, Diabetes, Hypertension Effect of Biolgical Rhythm and Exercise on Gene Expression Insulin-mimetic Activity of Soy Isoflavone Hypocholesteroleamic Effect of Sulfur-Containing Amino Acid Anti-obesity Effect of Dietary Amino Acid Supplementation
Stimulation of Glucose Uptake by a Soy Isoflavone, Genistein, in L6 Myotubes 【Cells】 L6 myotubes (from rattus) Differentiation for 【Methods】 Cultivation for 4 hours nine to eleven days Precultivation Stimulation by 50 μ M genistein Cultivation in DME medium for 2 hours DNA Microarray and Pathyway Analysis
Genistein Seems to Activate Two Pathways Leading to GLUT4 Translocation Exercise Insulin The rise of the ratio of AMP to ATP Insulin receptor Activation of some protein kinases The activation of AMPK including IRS-1 GLUT4 GLUT4 VAMP2 Translocation VAMP7 Translocation Glucose Glucose
A Typical Flow of DNA Microarray Data Analysis for Food and Nutrition Science Raw Data Normalization Filtering Cluster Analysis, Principal Component Analysis, etc. （ Tendency, Extraction of Marker Genes ） Each Responder Genes （ Rank Analysis, Statistical, Fold change…) Annotation Interpretation Functional Category Discussion Pathway Analysis Comparison with Other Data, Database 20
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