Document Type : Original Article

Authors

1 Department of Biochemistry, Universiti Kebangsaan Malaysia Medical Center, Jalan Yaacob Latif, 56000 Kuala Lumpur, Malaysia

2 Department of Biochemistry, Universiti Kebangsaan Malaysia Medical Center, Kuala Lumpur, Malaysia

Abstract

Objective: Oxidative stress that damages proteins result in aging and age related diseases. The aim of this study is to determine the effect of tocotrienol rich fraction (TRF) on the expression of proteins in oxidative stress-induced caenohabditis elegans (C.elegans) which has homologous genes to humans. Methods: The worms were treated with TRF prior to, after and continuously in separate groups upon induction of oxidative stress with hydrogen peroxide. The expression of proteins were analyzed with 2D-gel electrophoresis and identified with mass spectrophotometry. Results showed that induction of oxidative stress and TRF treatment separately modulated the expression of 11 proteins. Pre-treatment of TRF altered the expression of 5 proteins while post-treatment and a continuous treatment of TRF in oxidative stress-induced worms affected the expression of 9 and 10 proteins respectively. Results: From these differentially expressed proteins, a total of 12 were successfully identified. TRF was found to increase the expression of glutathione-S transferase but decreased the expression of mRNA cap guanine-N7 methyltransferase, inorganic pyro-phosphatase, enoyle-CoA hydratase, vitellogenin 6 precursor, cathepsin B-like cysteine proteinase 4 precursor, triosephosphatase isomerase, tubulin-specific chaperon B and putative D-amino acid oxidase. In conclusion, TRF modulated the expression of proteins involved in energy metabolism, oxidative stress, proteolysis and biosynthesis of mRNA in C.elegans.

Keywords

Main Subjects

Agnihotri G, Liu HW (2003). Enoyl-Coa Hydratase. Reaction, Mechanism, and Inhibition. Bioorg Med Chem 11(1): 9-20.
Avarre JC, Lubzens E, Babin PJ (2007). Apolipocrustacein, Formerly Vitellogenin, Is the Major Egg Yolk Precursor Protein in Decapod Crustaceans and Is Homologous to Insect Apolipophorin Ii/I and Vertebrate Apolipoprotein B. BMC Evol Biol 7(1): 3.
Berbée M, Fu Q, Boerma M, Kumar KS, Loose DS, HauerJensen M (2012). Mechanisms Underlying the Radioprotective Properties of Γ-Tocotrienol:
Comparative Gene Expression Profiling in Tocol-Treated Endothelial Cells. Genes Nutr 7(1): 75-81.
Brenner S (1974). The Genetics of Caenorhabditis Elegans. Genetics 77(1): 71-94.
Britton C, Murray L (2004). Cathepsin L Protease (Cpl-1) Is Essential for Yolk Processing During Embryogenesis in Caenorhabditis Elegans. J Cell Sci 117(21): 5133-5143.
Budin SB, Othman F, Louis SR, Bakar MA, Das S, Mohamed J (2009). The Effects of Palm Oil TocotrienolRich Fraction Supplementation on Biochemical
Parameters, Oxidative Stress and the Vascular Wall of Streptozotocin-Induced Diabetic Rats. Clinics 64(3):235-244.
Chin SF, Ibrahim J, Makpol S, Abdul Hamid NA, Abdul Latiff A, Zakaria Z, Mazlan M, Mohd Yusof YA, Karim AA, Wan Ngah WZ (2011). Tocotrienol Rich Fraction Supplementation Improved Lipid Profile and Oxidative
Status in Healthy Older Adults: A Randomized Controlled Study. Nutr Metab 8(42): 1-14.
Compagno C, Brambilla L, Capitanio D, Boschi F, Ranzi BM, Porro D (2001). Alterations of the Glucose Metabolism in a Triose Phosphate Isomerase-Negative Saccharomyces Cerevisiae Mutant. Yeast 18(7): 663-
Cox-Paulson EA, Grana TM, Harris MA, Batzli JM (2012). Studying Human Disease Genes in Caenorhabditis Elegans: A Molecular Genetics Laboratory Project. CBELife Sciences Education 11(2): 165-179.
Cutter AD, Dey A, Murray RL (2009). Evolution of the Caenorhabditis Elegans Genome. Mol Biol Evol 26(6): 1199-1234.
Dalton JP, Neill SO, Stack C, Collins P, Walshe A, Sekiya M, Doyle S, Mulcahy G, Hoyle D, Khaznadji E, Moiré N, Brennan G, Mousley A, Kreshchenko N, Maule AG, Donnelly SM (2003). Fasciola Hepatica Cathepsin L-Like Proteases: Biology, Function, and Potential in the Development of First Generation Liver Fluke Vaccines. Int J Parasitol 33(11): 1173-1181.
Delcroix M, Sajid M, Caffrey CR, Lim KC, Dvořák J, Hsieh I, Bahgat M, Dissous C, Mckerrow JH (2006). A Multienzyme Network Functions in Intestinal Protein
Digestion by a Platyhelminth Parasite. J Biol Chem 281(51): 39316-39329.
Depina AS, Iser WB, Park SS, Maudsley S, Wilson MA, Wolkow CA (2011). Regulation of Caenorhabditis Elegans Vitellogenesis by Daf-2/Iis through Separable Transcriptional and Posttranscriptional Mechanisms. BMC Physiol 11(1): 11.
Fabrega C, Hausmann S, Shen V, Shuman S, Lima CD (2004). Structure and Mechanism of Mrna Cap (Guanine-N7) Methyltransferase. Mol Cell 13(1): 77-89.
Fischer M, Regitz C, Kull R, Boll M, Wenzei U (2013). Vitellogenins Increase Stress Resistance of Caenorhabditis Elegans after Photorhabdus Luminescens
Infection Depending on the Steroid-Signaling Pathway. Microb Infect 15(8-9): 569-578.
Fusco D, Colloca G, Monaco MRL, Cesari M (2007). Effects of Antioxidant Supplementation on the Aging Process. Clin Interv Aging 2(3): 377.
Gao X, Albena TDK, Talalay P (2001). Powerful and Prolonged Protection of Human Retinal Pigment Epithelial Cells, Keratinocytes, and Mouse Leukemia Cells against Oxidative Damage: The Indirect Antioxidant
Effects of Sulforaphane. Proc Natl Acad Sci 98(26): 15221-15226.
Goon JA, Zainudin MS, Karim NA, Wan Ngah WZ (2013). Effect of the Tocotrienol-Rich Fraction on the Lifespan and Oxidative Biomarkers in Caenorhabditis Elegans under Oxidative Stress. Clinics 68(5): 599-604.
Grant CM (2008). Metabolic Reconfiguration Is a Regulated Response to Oxidative Stress. J Biol 7(1): 1. 
Guiliano DB, Hong X, Mckerrow JH, Blaxter ML, Oksov Y, Liu J, Ghedin E, Lustigman S (2004). A Gene Family of Cathepsin L-Like Proteases of Filarial Nematodes Are Associated with Larval Molting and Cuticle and Eggshell
Remodeling. Mol Biochem Parasitol 136(2): 227-242.
Hashmi S, Zhang J, Oksov Y, Lustigman S (2004). The Caenorhabditis Elegans Cathepsin Z-Like Cysteine Protease, Ce-Cpz-1, Has a Multifunctional Role During the Worms' Development. J Biol Chem 279(7): 6035-6045.
Hayes JD, Flanagan JU, Jowsey IR (2005). Glutathione Transferases. Ann Rev Pharmacol Toxicol 45(1): 51-88.
Johansson AS, Mannervik B (2001). Human Glutathione Transferase A3-3, a Highly Efficient Catalyst of DoubleBond Isomerization in the Biosynthetic Pathway of Steroid Hormones. J Biol Chem 276(35): 33061-33065.
Jowsey IR, Thomson AM, Flanagan JU, Murdock PR,
Moore GBT, Meyer DJ, Murphy GJ, Smith SA, Hayes JD (2001). Mammalian Class Sigma Glutathione STransferases: Catalytic Properties and Tissue-Specific Expression of Human and Rat Gsh-Dependent
Prostaglandin D2 Synthases. Biochem J 359(507-516. 
Kannappan R, Gupta SC, Kim JH, Aggarwal BB (2012). Tocotrienols Fight Cancer by Targeting Multiple Cell Signaling Pathways. Genes Nutr 7(1): 43-52.
Ko KM, Lee W, Yu JR, Ahnn J (2007). Pyp-1, Inorganic Pyrophosphatase, Is Required for Larval Development and Intestinal Function in C. Elegans. FEBS letters 581(28): 5445-5453.
Kunwar A, Jayakumar S, Bhilwade HN, Bag PP, Bhatt H, Chaubey RC, Priyadarsini KI (2011). Protective Effects of Selenocystine against Γ-Radiation-Induced Genotoxicity in Swiss Albino Mice. Radiat Environ
Biophys 50(2): 271-280.
Larsen PL (1993). Aging and Resistance to Oxidative Damage in Caenorhabditis Elegans. Proc Natl Acad Sci U S A 90(19): 8905-8909.
Lee OH, Kwon YI, Hong HD, Park CS, Lee BY, Kim YC (2009). Production of Reactive Oxygen Species and Changes in Antioxidant Enzyme Activities During
Differentiation of 3t3-L1 Adipocyte. J Korean Soc Appl Biol Chem 52(1): 70-75.
Lindblom TH, Dodd AK (2006). Xenobiotic Detoxification in the Nematode Caenorhabditis Elegans. J Exp Zool A Comp Exp Biol 305(9): 720-730.
Lopez-Fanarraga M, Carranza G, Bellido J, Kortazar D, Villegas JC, Zabala JC (2007). Tubulin Cofactor B Plays a Role in the Neuronal Growth Cone. J Neurochem 100(6):1680-1687.
Makpol S, Abidin AZ, Sairin K, Mazlan M, Top GM, Wan Ngah WZ (2010). Γ-Tocotrienol Prevents Oxidative229 Goon et al / Int. J. Adv. Biol. Biom. Res, 2015; 3 (3), 222-230
Stress-Induced Telomere Shortening in Human Fibroblasts Derived from Different Aged Individuals. Oxid Med Cell Longev 3(1): 35-43.
Makpol S, Zainuddin A, Chua KH, Mohd Yusof YM, Wan Ngah WZ (2012). Gamma-Tocotrienol Modulation of Senescence-Associated Gene Expression Prevents Cellular Aging in Human Diploid Fibroblasts. Clinics 67(2): 135-143.
Markaki M, Tavernarakis N (2010). Modeling Human Diseases in Caenorhabditis Elegans. Biotech J 5(12):1261-1276.
Miranda-Miranda E, Zamora-Ruiz A, Cossio-Bayugar R (2009). Molecular Cloning and Expression of a Caenorhabditis Elegans Cathepsin B-Like Protease. Biotech 8(2): 242-247.
Mohamed MM, Sloane BF (2006). Cysteine Cathepsins: Multifunctional Enzymes in Cancer. Nat Rev Cancer 6(10): 764-775.
Muid S, Ali AM, Yusoff K, Nawawi H (2013). Optimal Antioxidant Activity with Moderate Concentrations of Tocotrienol Rich Fraction (Trf) in in Vitro Assays. Int Food Res J 20(2): 687-694.
Nakamura A, Yasuda K, Adachi H, Sakurai Y, Ishii N, Goto S (1999). Vitellogenin-6 Is a Major Carbonylated Protein in Aged Nematode, Caenorhabditis Elegans. Biochem Biophys Res Commun 264(2): 580-583.
Nakamura H, Fang J, Maeda H (2012). Protective Role of D-Amino Acid Oxidase against Staphylococcus Aureus Infection. Infect Immun 80(4): 1546-1553.
Nazrun AS, Khairunnur A, Norliza M, Norazlina M, Nirwana, I (2008). Effects of Palm Tocotrienols on Oxidative Stress and Bone Strength in Ovariectomised
Rats. Med Health 3(2): 247-255.
Nguyen T, Sherratt PJ, Pickett CB (2003). Regulatory Mechanisms Controlling Gene Expression Mediated by the Antioxidant Response Element. Ann Rev Pharmacol Toxicol 43(1): 233-260.
Orosz F, Oláh J, Ovádi J (2006). Triosephosphate Isomerase Deficiency: Facts and Doubts. IUBMB life 58(12): 703-715.
Requena JR, Chaos CC, Levine RL, Stadtman ER (2001). Glutamic and Aminoadipic Semialdehydes Are the Main Carbonyl Products of Metal-Catalyzed Oxidation of Proteins. Proc Natl Acad Sci 98(1): 69-74.
Requena JR, Stadtman ER (1999). Conversion of Lysine to Nϵ-(Carboxymethyl)Lysine Increases Susceptibility of Proteins to Metal-Catalyzed Oxidation. Biochem Biophys Res Comm 264(1): 207-211.
Saitoh Y, Katane M, Kawata T, Maeda K, Sakamoto T, Inoue T, Arai H, Nakagawa Y, Homma H (2012). Spatiotemporal Localization of D-Amino Acid Oxidase and D-Aspartate Oxidases During Development in
Caenorhabditis Elegans. Mol Cell Biol 32(10): 1967- 1983.
Shevchenko A, Wilm M, Vorm O, Mann M (1996). Mass Spectrometric Sequencing of Proteins Silver-Stained Polyacrylamide Gels. Anal Chem 68(5): 850-858.
Shukla V, Yadav D, Phulara SC, Gupta MM, Saikia SK, Pandey R (2012). Longevity-Promoting Effects of 4- Hydroxy-E-Globularinin in Caenorhabditis Elegans. Free Rad Biol Med 53(10): 1848-1856. Srayko M, Kaya A, Stamford J, Hyman AA (2005). Identification and Characterization of Factors Required
for Microtubule Growth and Nucleation in the Early C. Elegans Embryo. Dev Cell 9(2): 223-236.
Stiernagle T (2006). Maintenance of C. Elegans. WormBook 1-11.
Symersky J, Li S, Carson M, Luo M (2003). Structural Genomics of Caenorhabditis Elegans: Triosephosphate Isomerase. Proteins: Structure, Function, and Bioinformatics 51(3): 484-486.
Szolajska E, Chroboczek J (2011). Faithful Chaperones. Cell Mol Life Sci 68(20): 3307-3322.
Tedeschi G, Pollegioni L, Negri A (2012). Assays of DAmino Acid Oxidases. In (edit). Unnatural Amino Acids 381-395. Springer.
Then SM., Sanfeliu C, Top GM, Wan Ngah WZ, Mazlan M (2012). Gamma-Tocotrienol Does Not Substantially Protect Ds Neurons from Hydrogen Peroxide-Induced Oxidative Injury. Nutr Metab 9(1): 1-24.
Williamson AL, Brindley PJ, Knox DP, Hotez PJ, Loukas A (2003). Digestive Proteases of Blood-Feeding Nematodes. Trends Parasitol 19(9): 417-423.
Williamson AL, Lecchi P, Turk BE, Choe Y, Hotez PJ, Mckerrow JH, Cantley LC, Sajid M, Craik CS, Loukas A (2004). A Multi-Enzyme Cascade of Hemoglobin
Proteolysis in the Intestine of Blood-Feeding Hookworms. J Biol Chem 279(34): 35950-35957.
Wilm M, Shevchenko A, Houthaeve T, Breit S, Schweigerer L, Fotsis T, Mann M (1996). Femtomole Sequencing of Proteins from Polyacrylamide Gels by
Nano-Electrospray Mass Spectrometry. Nature 379(6564): 466-469.
Yan, J. X., Wait, R., Berkelman, T., Harry, R. A., Westbrook, J. A., Wheeler, C. H. & Dunn, M. J. 2000. A Modified Silver Staining Protocol for Visualization of Proteins Compatible with Matrix-Assisted Laser Desorption/Ionization and Electrospray Ionization-Mass Spectrometry. Electrophoresis 21(17): 3666-3672.
Yi, Y.-J., Sutovsky, M., Kennedy, C. & Sutovsky, P. 2012. Identification of the Inorganic Pyrophosphate Metabolizing, Atp Substituting Pathway in Mammalian Spermatozoa PloS one 7(4): e34524.
Zheng, S. & Shuman, S. 2008. Mutational Analysis of Vaccinia Virus Mrna Cap (Guanine-N7) Methyltransferase Reveals Essential Contributions of the N-Terminal Peptide That Closes over the Active Site Rna 14(11):2297-2304.