Proteomic analysis of ‘Zaosu’ pear (Pyrus bretschneideri Rehd.) and its red skin bud mutation

Proteome Science

Junke Zhang¹  Lingfei Xu¹  Peng Zhou¹  Zonghao Qiu¹  Min Hu¹ 

[1] Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest China, Ministry of Agriculture, Yangling, Shaanxi, 712100, China

关键词: MALDI TOF/TOF-MS;    Plant proteomics;    Red skin bud mutation;    Pear;   

Others  :  817202
DOI  :  10.1186/1477-5956-10-51

 received in 2012-02-17, accepted in 2012-07-17,  发布年份 2012

【 摘 要 】

Background

Breeding for strong red skin color is an important objective of the pear breeding program. There are few reports of proteome research in green skin pear and its red skin bud mutation. The manuscript at hand is one of the first studies dealing with 2D-PAGE-based analysis of pear fruits and leaves, establishing a suitable sample preparation and testing different 2D-PAGE protocols. Therefore, it may grant a basis for further studies on the pear proteome being the studies main goal. A proteomic analysis was conducted on leaves and fruits of ‘Zaosu’ pear (Pyrus bretschneideri Rehd.) and its red skin bud mutation in order to reveal their genetic differences in the protein level.

Results

In the present study, the optimized two-dimensional (2-D) gel electrophoresis system of pear leaf and fruit was set up, and applied to analyze the leaves and fruit protein. The interesting peptide fragments were determined using 4800 Plus MALDI TOF/TOFTM Analyzer mass spectrometer, and the sequence obtained was blasted in NCBInr to identify the differentially-expressed protein. In the 1.5-fold differently-expressed proteins between ‘Zaosu’ pear and its mutant, 10 out of 35 proteins in fruit and 12 out of 24 ones in leaves were identified successfully. Among the 22 identified proteins, 7 protein spots were related to photosynthesis and energy metabolism; 4 were associated with environmental stress; 4 with disease defense; 2 with amino acid metabolism; 2 with cytoskeleton; 1 with antioxidant function; 1 with calcium metabolism; and 1 with unknown function. Moreover, related physiological index, such as chlorophyll content, Rubisco content and polyphone oxidase activity, were different between ‘Zaosu’ pear and its mutant.

Conclusion

A 2-D gel electrophoresis system of pear leaves and fruits was established, which was suitable for the analysis of proteome comparison. To the best of our knowledge, we have performed the first analysis of the proteomic changes in leaves and fruits of ‘Zaosu’ pear and its red skin bud mutation. Our study provides important information on the use of proteomic methods for studying protein regulation of ‘Zaosu’ pear and its red skin bud mutation.

【 授权许可】

   
2012 Hu et al; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

【 参考文献 】

• [1]Volz RK, White AG, Brewer LR: Breeding for red skin color in interspecific pears. Acta Hort 2008, 800:469-474.
• [2]Xu LF: A fine line of precocity red pear. Shanxi Fruits 2009, 4:44-45.
• [3]Prinsi B, Negri AS, Fedeli C, Morgutti S, Negrini N, Cocucci M, Espen L: Peach fruit ripening: A proteomic comparative analysis of the mesocarp of two cultivars with different flesh firmness at two ripening stages. Phytochemistry 2011, 72:1251-1262.
• [4]Muccilli V, Licciardello C, Fontanini D, Russo MP, Cunsolo V, Saletti R, Recupero GR, Foti S: Proteome analysis of Citrus Sinensis L. (Osbeck) flesh at ripening time. J Proteomics 2009, 73:134-52.
• [5]Feng S, Chen X, Zhang Y, Wang Y, Song Y, Chen XL, Li X, Li M, Liu J, Wang Q, Liu M: Differential expression of proteins in red pear following fruit bagging treatment. Protein J 2011, 30:194-200.
• [6]Negri AS, Prinsi B, Rossoni M, Failla O, Scienza A, Cocucci M, Espen L: Proteome changes in the skin of the grape cultivar Barbera among different stages of ripening. BMC Genomics 2008, 9:378. BioMed Central Full Text
• [7]Martinez-Esteso MJ, Sellés-Marchart S, Vera-Urbina JC, Pedreño MA, Bru-Martinez R: Changes of defense proteins in the extracellular proteome of grapevine (Vitis vinifera cv. Gamay) cell cultures in response to elicitors. Proteomics 2009, 73:331-41.
• [8]Hartman DJ, Dougan D, Hoogenraad NJ, Høj PB: Heat shock proteins of barley mitochondria and chloroplasts. Identification of organellar hsp 10 and 12: putative chaperonin 10 homologues. FEBS Letters 1992, 305:147-150.
• [9]Lund AA, Blum PH, Bhattramakki D, Elthon T: Heat-stress response of maize mitochondria. Plant Physiol 1998, 116:1097-1110.
• [10]Sanmiya K, Suzuki K, Egawa Y: Mitochondrial small heat shock protein enhances thermotolerance in tobacco plants. FEBS Lett 2004, 557:265-268.
• [11]Klimmek F, Sjödin A, Noutsos C, Leister D, Jansson S: Abundantly and rarely expressed Lhc protein genes exhibit distinct regulation patterns in plants. Plant Physiol 2006, 140:793-804.
• [12]Luciński R, Jackowski G: The structure, functions and degradation of pigment-binding proteins of photosystem II. Acta Biochim Pol 2006, 53:693-708.
• [13]Liu C, Zhang Y, Cao D: Structural and functional analysis of the antiparallel strands in the lumenal loop of the major light-harvesting chlorophyll a/b complex of photosystem II (LHCIIb) by site-directed mutagenesis. J of Biological Chemistry 2008, 283:487-495.
• [14]Teramoto H, Ono T, Minagawa J: Identification of Lhcb gene family encoding the light harvesting chlorophyll a/b proteins of photosystem II in Chlamydomonas reinhardtii. Plant Cell Physiol 2001, 42:849-856.
• [15]Tokutsu R, Teramoto H, Takahashi Y, Ono T, Minagawa J: The light-harvesting complex of photosystem I in Chlamydomonas reinhardtii: protein composition, gene structures and phylogenic implications. Plant Cell Physiol 2004, 45:138-145.
• [16]Chu HA, Nguyen AP, Debus RJ: Amino Acid Residues That Influence the Binding of Manganese or Calcium to Photosystem II. 1. The Lumenal Interhelical Domains of the D1 Polypeptide. Biochemistry 1995, 34:5839-5858.
• [17]Tang XS, Diner BA, Larsen BS: Identification of histidine at the catalytic site of the photosynthetic oxygen evolving complex. Proc Natl Acad Sci 1994, 91:704-708.
• [18]Magnuson A, Andreasson LE: Different manganese binding sites in Photosystem II probed by selective chemical modification of histidyl and carboxylic acid residues. Biochemistry 1997, 36:3254-3261.
• [19]Ghirardi ML, Preston C, Seibert M: Use of a novel histidyl modifier to probe for residues on tris treated photosystem II membrane fragments that may bind functional manganese. Biochemistry 1998, 37:13567-13574.
• [20]Makino A, Mae T, Ohira K: Photosynthesis and ribulose 1,5-bisphosphate carboxylase in rice leaves: changes in photosynthesis and enzymes involved in carbon assimilation from leaf development through senescence. Plant Physiology 1983, 73:1002-1007.
• [21]To KY, Suen DF, Grace-Chen SC: Molecular characterization of ribulose-1,5-bisph.osphate carboxylase/oxygenase activase in rice leaves. Planta 1999, 209:66-76.
• [22]Portis AR: Rubisco activase – Rubisco's catalytic chaperone. Photosynth Res 2003, 75:11-27.
• [23]Touchette BW, Burkholder JM: Overview of the physiological ecology of carbon metabolism in seagrasses. J Exp Mar Bio Ecol 2000, 250:169-205.
• [24]Gabai VL, Kabakov AE: Rise in heat shock protein level confers tolerance to energy deprivation. FEBS Lett 1993, 327:247-250.
• [25]Banti V, Loreti E, Novi G: Heat acclimation and cross-tolerance against anoxia in Arabidopsis. Plant Cell Environ 2008, 31:1029-1037.
• [26]Downs CA, Heckathorn SA: The mitochondrial small heat shock protein protects NADH: ubiquinone oxidoreductase of the electron transport chain during heat stress in plants. FEBS lett 1998, 430:246-250.
• [27]Gupta RS: Phylogenetic analysis of the 90 KD heat shock family of protein sequences and an examination of the relationship among animals, plants, and fungi species. Mol Biol Evol 1995, 12:1063-1073.
• [28]Lüders J, Demand J, Höhfeld J: The ubiquitin-related BAG-1 provides a link between the molecular chaperones Hsc70/Hsp70 and the proteasome. J Biol Chem 2000, 275:4613-4617.
• [29]Lin YZ, Chen HY, Kao R, Chang SP, Chang SJ, Lai EM: Proteomic analysis of rice defense response induced by probenazole. Phytochemistry 2008, 69:715-728.
• [30]Baluska F, Hlavacka A, Samaj J, Palme K, Robinson DG, Matoh T, McCurdy DW, Menzel D, Volkmann D: F-actin-dependent endocytosis of cell wall pectins in meristematic root cells. Insights from brefeldin A-induced compartments. Plant Physiol 2002, 130:422-431.
• [31]Starr DA, Han M: An Chors away: an actin based mechanism of nuclear positioning. J Cell Sci 2003, 116:211-216.
• [32]Yang SF, Hoffman NE: Ethylene biosynthesis and its regulation in higher plants. Annu Rev Plant Physiology 1984, 35:155-189.
• [33]Heby O, Persson L: Molecular genetics of polyamine synthesis in eukaryotic cells. Trends in Biochemical Sci 1990, 15:153-158.
• [34]Chen H, Xiong L: Pyridoxine is required for post-embryonic root development and tolerance to osmotic and oxidative stresses. Plant J 2005, 44:396-408.
• [35]Wang H, Liu D, Liu C: The pyridoxal kinase gene TaPdxK from wheat complements vitamin B6 synthesis-defective Escherichia coli. J Plant Physiology 2004, 161:1053-1060.
• [36]Ehrenshaft M, Bilski P, Li M: A highly conserved sequence is a novel gene involved in de novo vitamin B6 biosynthesis. Microbiology 1999, 96:9374-9378.
• [37]Gallardo K, Job C, Groot SPC, Puype M, Demol H, Vandekerckhove J, Job D: Proteomic analysis of Arabidopsis seed germination of priming. Plant Physiol 2001, 126:835-848.
• [38]Bradford MM: A rapid and sensitive method for quantitation of microgram quantities of protein utilizing principle of protein-dye binding. Anal Biochem 1976, 72:248-254.
• [39]Candiano G, Bruschi M, Musante L, Santucci L, Ghiggeri GM, Carnemolla B, Orecchia P, Zardi L, Righetti PG: Blue silver: A very sensitive colloidal Coomassie G-250 staining for proteome analysis. Electrophoresis 2004, 25:1327-1333.
• [40]Gao JF: Measurement of chlorophyll content. In The guidance of plant physiology experiments. 1st edition. Edited by Gao JF. Xi’an: Higher education press; 2006:74-76.
• [41]Lilley RM, Walker DA: An improved spectrophotometric assay for ribulose bisphosphate carboxylase. Biochemica et biophysica Acta 1974, 358:226-229.
• [42]Vaughn KC, Lax AR, Duke SO: Polyphenol oxidase: The chloroplast oxidase with no established function. Physiol Plant 1988, 72:659-665.