巴豆醛对水稻悬浮培养细胞蛋白质表达的影响

摘要/Abstract

摘要： 对巴豆醛处理水稻悬浮培养细胞后蛋白质组水平的变化进行了研究,以期发现在调控水稻细胞周期活动中起重要作用的蛋白。采用双向电泳分离蛋白,PDQuest软件分析凝胶图谱确定差异蛋白点,LCQ-Fleet ion trap系统鉴定蛋白,相关数据库检索分析差异蛋白点,获得了45个差异蛋白点,经质谱鉴定和数据库分析得到26个可信蛋白,集中在胁迫反应、蛋白质命运,信号转导,物质与能量代谢和膜功能等。在这些蛋白中,小分子量热激蛋白、Skp1蛋白和26S蛋白酶,CDC48,6-磷酸葡萄糖胺乙酰基转移酶以及蔗糖合酶等蛋白已有文献报告其在植物细胞周期调控中的作用,其它一些鉴定的蛋白参与了细胞的生理活动,其与细胞周期间的相互关系需进一步的研究。巴豆醛处理影响了水稻悬浮培养细胞正常的细胞周期活动,鉴定出部分蛋白与细胞周期的调控具有密切关系,而一些与水稻细胞分化、发育相关蛋白也得到了鉴定,其在细胞周期上的作用尚未有文献报告,这些蛋白的进一步研究将有助于揭示水稻细胞周期的调控机制。

关键词: 巴豆醛, 水稻悬浮培养细胞, 差异蛋白质组, 细胞周期

Abstract: In order to study the important proteins related to cell cycle of rice, the effect of crotonaldehyde on the protein expression of suspension cell line of rice was investigated. The proteins were separated by the method of two dimensional electrophoresis, the different protein spots were analyzed by PDQuest software, and then the different protein spots were identified by LCQ-Fleet ion trap and database searching. The results showed that there were forty-five protein spots expressed differently, and twenty-six of them were identified by mass spectrometry and database analysis. The predicted function of these proteins was related to stress response, protein synthesis and processing, signal transduction, metabolism membrane function. Some of the proteins, including small molecular proteins, Skp1, 26S proteasome, cell division cycle protein 48, glucosamine-6-phosphate acetyltransferase and sucrose synthase, were related with the cell cycle process. Other identified proteins should be study further to explore its function in cell cycle of rice. The normal cell cycle activity was effected by crotonaldehyde, some of the identified proteins were related with regulation of cell cycle. Other proteins related to cell differentiation had no reports about their function to cell cycle;these proteins should be study further to explore the mechanism of cell cycle regulation.

Key words: Crotonaldehyde, Rice suspension cells, Different proteomics, Cell cycle

刘悦萍, 郭蓓, 胡昊, 张国庆. 巴豆醛对水稻悬浮培养细胞蛋白质表达的影响[J]. 生物技术通报, 2014, 0(10): 119-127.

Liu Yueping, Guo Bei, Hu Hao, Zhang Guoqing. Effects of Crotonaldehyde on the Protein Expression of Rice Suspension Cells[J]. Biotechnology Bulletin, 2014, 0(10): 119-127.

参考文献

[1] Vandepoele K, Raes J, De Veylder L, et al. Genome-wide analysis of core cell cycle genes in Arabidopsis[J]. Plant Cell, 2002, 14：903-916.
[2] Wang G, Kong H, Sun Y, et al. Genome-wide analysis of the cyclin family in Arabidopsis and comparative phylogenetic analysis of plant cyclin-like proteins[J]. Plant Physiol, 2004, 135：1084-1099.
[3] Menges M, Hennig L, Gruissem W, et al. Cell cycle-regulated gene expression in Arabidopsis[J]. J Biol Chem, 2002, 77：41987-42002.
[4] Zhao XA, Harashima H, Dissmeyer N, et al. A general G1/S-phase cell-cycle control module in the flowering plant Arabidopsis thaliana[J]. PLoS Genetics, 2012, 8（8）：e1002847.
[5] Leene JV, Hollunder J, Eeckhout D, et al. Targeted interactomics rev-eals a complex core cell cycle machinery in Arabidopsis thaliana[J]. Molecular Systems Biology, 2010, 6：397-405.
[6] Guo J, Song J, Wang F, et al. Genome-wide identification and expression analysis of rice cell cycle genes[J]. Plant Molecular Biology, 2007, 64（4）：349-360.
[7] Dori-Bachash M, Dassa B, Pietrokovski S, et al. Proteome-based comparative analyses of growth stages reveal new cell cycle-dependent functions in the predatory bacterium Bdellovibrio bacteriovorus[J]. Applied and Environmental Microbiology, 2008：7152-7162.
[8] Agrawal GK, Rakwal R. Rice proteomics：A move toward expanded proteome coverage to comparative and functional proteomics uncovers the mysteries of rice and plant biology[J]. Proteomics, 2011, 11（9）：1630-1649.
[9] Neudecker T, Eder E, Deininger C, et al. Crotonaldehyde is mutagenic in Salmonella typhimurium TA100[J]. Environmental Molecular Mutagen, 1989, 14：146-148.
[10] Czerny C, Eder E, Runger TM. Genotoxicity and mutagenicity of the a, b-unsaturated carbonyl compound crotonaldehyde（butenal）on a plasmid shuttle vector[J]. Mutation Research, 1998, 407：125-134.
[11] Fernandes PH, Kanuri M, Nechev LV, et al. Mammalian cell muta-genesis of the DNA adducts of vinyl chloride and crotonaldehyde[J]. Environmental Molecular Mutagen, 2005, 45：455-459.
[12] Jenes B, Pauk J. Plant regeneration from protoplast derived calli in rice（Oryza sativa L.）using dicamba[J]. Plant Science, 1989, 62（3）：187-198.
[13] Wessel D, Flügge UI. A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids[J]. Analytical Biochemistry, 1984, 138（1）：141-143.
[14] 刘悦萍, 杨爱珍, 郭蓓, 等.适合水稻悬浮培养细胞蛋白质组分析的双向电泳技术[J].生物技术通报, 2010（11）：82-86.
[15] Bryant JA, Francis D. The plant cell cycle[J]. Annals of Botany, 2011, 107：1063.
[16] Liu XY, Yang ZH, Pan XJ, et al. Gene expression profile and cytotoxicity of human bronchial epithelial cells exposed to crotonaldehyde[J]. Toxicology Letters, 2010a, 197（2）：113-122.
[17] Liu XY, Yang ZH, Pan XJ, et al. Crotonaldehyde induces oxidative stress and caspase-dependent apoptosis in human bronchial epithelial cells[J]. Toxicology Letters, 2010b, 195（1）：90-98.
[18] ER, Lee GJ, Vierling E. Evolution, structure and function of the small heat shock proteins in plants[J]. Journal of Experimental Botany, 1996, 47（3）：325-338.
[19] Mehlen P, Schulze-Osthoff K, Arrigo AP. Small stress proteins as novel regulators of apoptosis[J]. Journal of Biological Chemistry, 1996, 271（28）：16510-16514.
[20] Kamradt MC, Chen F, Sam S. The small heat shock protein alpha B-crystallin negatively regulates apoptosis during myogenic differentiation by inhibiting caspase-3 activation[J]. Journal of Biological Chemistry, 2002, 277（41）：38731-38736.
[21] Liu S, Li J, Tao Y, et al. Small heat shock protein alpha Bcrystallin binds to p53 to sequester its translocation to mitochondria during hydrogen peroxide-induced apoptosis[J]. Biochemical Biophysical Research Communications, 2007, 354（1）：109-114.
[22] J, Vierstra RD. The ubiquitin 26S proteasome proteolytic pathway[J]. Annual Review Plant Biology, 2004, 55：555-590.
[23] W, Walz J, Zühl F, Seemüller E. The proteasome：Paradigm of a self compartmentalizing protease[J]. Cell, 1998, 92：367-380.
[24] R, Lu P, O’Neil SD. Arabidopsis SKP1, a homologue of a cell cycle regulator gene, is predominantly expressed in meriste-matic cells[J]. Planta, 1998, 204（3）：345-351.
[25] Vierstra RD. The ubiquitin-26S proteasome system at the nexus of plant biology[J]. Nature Reviews Molecular Cell Biology, 2009, 10：385-397.
[26] Ciechanover A, Orian A, Schwartz AL. Ubiquitin-mediated proteol-ysis：biological regulation via destruction[J]. Bioessays, 2000, 22：442-451.
[27] CM. Mechanisms underlying ubiquitination[J]. Annual Review Biochemistry, 2001, 70：503-533.
[28] HS, Desprez T, Santoni V, et al. The higher plant Arabidopsis thaliana encodes a functional CDC48 homologue which is highly expressed in dividing and expanding cells[J]. The EMBO Journal, 1995, 14：5626-5637.
[29] Fröhlich KU, Fries HW, Rudiger M, et al. Yeast cell cycle protein CDC48p shows full-length homology to the mammalian protein VCP and is a member of a protein family involved in secretion, peroxisome formation, and gene expression[J]. Journal of Cell Biology, 1991, 114：443-453.
[30] Shirogane T, Fukada T, Muller JM, et al. Synergistic roles for Pim-1 and c-Myc in STAT3-mediated cell cycle progression and Antiapoptosis[J]. Immunity, 1999, 11：709-719.
[31] Müller JM, Deinhardt K, Rosewell L, et al. Targeted deletion of p97（VCP/CDC48）in mouse resultsin early embryonic lethality[J]. Biochemical and Biophysical Research Communications, 2007, 354：459-465.
[32] Sookhee P, David MR, Sebastian YB. In planta analysis of the cell cycle-dependent localization of AtCDC48A and its critical roles in cell division, expansion, and differentiation[J]. Plant Physiology, 2008, 148：246-258.
[33] Hansol B, Soo MC, Seong WY. Suppression of the ER-localized AAA ATPase NgCDC48 inhibits tobacco growth and development[J]. Molecular Cells, 2009, 28：57-65.
[34] Ralf JB, Hans Z. Mechanisms of Cdc48/VCP-mediated cell death-from yeast apoptosis to human disease[J]. Biochimica et Biophy-sica Acta, 2008, 1783：1418-1435.
[35] Rati V, Robert O, Ruihua F, et al. Cdc48/p97 mediates UV-dependent turnover of RNA pol II[J]. Molecular Cell, 2010, 41：82-92.
[36] Yeung HO, Kloppsteck P, Niwa H, et al. Insights into adaptor bin-ding to the AAA protein[J]. Biochemical Society Transactions, 2008, 36：62-72.
[37] Müssig C, Fischer S, Altmann T. Brassinosteroid-regulated gene expression[J]. Plant Physiology, 2002, 129（7）：1241-1251.
[38] Mehra A, Wrana JL. TGF-β and the Smad signal transduction pathway[J]. Biochemistry and Cell Biology, 2002, 80（5）：605-622.
[39] Heike R, Herter T, Grishkovskaya I, et al. Crystal structure and functional characterization of a glucosamine-6-phosphate N-acetyl-transferase from Arabidopsis thaliana[J]. Biochemcal Journal, 2012, 443, 427-437.
[40] Nozakia M, Sugiyamab M, Duanc J, et al. A missense mutation in the glucosamine-6-phosphate N-Acetyltransferase-encoding gene causes temperature-dependent growth defects and ectopic lignin deposition in Arabidopsis[J]. The Plant Cell, 2012, 24（8）：3366-3379.
[41] Jiang H, Wang S, Dang L, et al. A novel short-root gene encodes a glucosamine-6-phosphate acetyltransferase required for maintaining normal root cell shape in rice[J]. Plant Physiology, 2005, 138：232-242.
[42] Riou-Khamlichi C, Menges M, Sandra HJM, et al. Sugar control of the plant cell cycle：differential regulation of Arabidopsis D-type cyclin gene expression[J]. Molecular and Cellular Biology, 2000, 7：4513-4521.
[43] Yang R, Tang Q, Wang H, et al. Analyses of two rice（Oryza sativa）cyclin-dependent kinase inhibitors and effects of transgenic expression of OsiICK6 on plant growth and development[J]. Annals of Botany, 2011, 107：1087-1101.
[44] Arpat AB, Waugh M, Sullivan JP, et al. Functional genomics of cell elongation in developing cotton fibers[J]. Plant Molecular Biology, 2004, 54：911-929.