F_1F_0-ATP合成酶β亚基与肿瘤相关性的初步研究
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摘要
过去认为F_1F_0-ATP合成酶只表达定位在线粒体膜上,但近年来越来越多的研究发现其亚基也分布在一些细胞的细胞质膜上,称为质膜异位F_1F_0-ATP合成酶。许多肿瘤细胞表面都存在F_1F_0-ATP合成酶,但是不同肿瘤细胞表面F_1F_0-ATP合成酶的量不同。F_1F_0-ATP合成酶β亚基的功能性单克隆抗体能够增强癌细胞及其耐药株对化疗药物的敏感性。本文旨在研究F_1F_0-ATP合成酶β亚基在肿瘤细胞中的定位及其表达量差异对细胞增殖的影响,进一步探讨肿瘤细胞膜表面表达F_1F_0-ATP合成酶的功能。
为研究F_1F_0-ATP合成酶β亚基在肿瘤细胞中的定位,我们利用荧光标记载体使β亚基带有荧光标签。从细胞中提取总RNA,反转录得到cDNA第一链,通过PCR扩增atp5b基因,克隆到真核表达载体pEGFP–N1中,转染细胞后,荧光显微镜下观察发现细胞表面确实有荧光存在。
为研究F_1F_0-ATP合成酶β亚基表达量差异对细胞增殖的影响,我们设计了3个shRNA片段干扰ATP5B蛋白的表达,转染细胞后分别在正常条件和缺氧条件下培养,MTT法检测细胞增殖能力,发现shRNA1、shRNA2组细胞增殖率大于1,shRNA3组细胞增殖率小于1。
F_1F_0-ATP synthase was considered as a protein strictly located in the inner membrane of mitochondria previously. However, current studies have demonstrated that components of F_1F_0-ATP synthase exist on the plasma membrane of tumor cells and adipocytes, which was published as ectopical plasma membrane F_1F_0-ATP synthase. F_1F_0-ATP synthase exist on the surface of many tumor cells, but always with different ratio. It is reported that the functional monoclonal antibody against theβsubunit of F_1F_0-ATP synthase can enhance the cancer and its resistant strains’sensitive to the chemotherapy. This research is aimed at studying the location of theβsubunit of F_1F_0-ATP synthase in tumor cells. At the same time, the effects of the different expression of theβsubunit to cell proliferation has been studied, and the function of F_1F_0-ATP synthase on the surface of tumor cell would be exploited.
In order to study the location of theβsubunit of F_1F_0-ATP synthase,βsubunit was cloned to the vector which contains Fluorescent labels. Extract the total RNA from cells, obtain cDNA by reverse transcript and use PCR to amplify atp5b, which was cloned to the Eukaryotic expression vector pEGFP–N_1. After the transfection, fluorescent was observed on cell surface under the fluorescence microscope.
To investigate the effects of the different expression of theβsubunit to cell proliferation, we designed three shRNA to interfered the expression of ATP5B. Cell was cultured on normal and hypoxic conditions after transfection, cell proliferation was detected by MTT assays. The results show the proliferation rate of cells transfected with shRNA1 or shRNA2 was more than 1, and cells transfected with shRNA3 was less than 1.
为研究F_1F_0-ATP合成酶β亚基在肿瘤细胞中的定位,我们利用荧光标记载体使β亚基带有荧光标签。从细胞中提取总RNA,反转录得到cDNA第一链,通过PCR扩增atp5b基因,克隆到真核表达载体pEGFP–N1中,转染细胞后,荧光显微镜下观察发现细胞表面确实有荧光存在。
为研究F_1F_0-ATP合成酶β亚基表达量差异对细胞增殖的影响,我们设计了3个shRNA片段干扰ATP5B蛋白的表达,转染细胞后分别在正常条件和缺氧条件下培养,MTT法检测细胞增殖能力,发现shRNA1、shRNA2组细胞增殖率大于1,shRNA3组细胞增殖率小于1。
F_1F_0-ATP synthase was considered as a protein strictly located in the inner membrane of mitochondria previously. However, current studies have demonstrated that components of F_1F_0-ATP synthase exist on the plasma membrane of tumor cells and adipocytes, which was published as ectopical plasma membrane F_1F_0-ATP synthase. F_1F_0-ATP synthase exist on the surface of many tumor cells, but always with different ratio. It is reported that the functional monoclonal antibody against theβsubunit of F_1F_0-ATP synthase can enhance the cancer and its resistant strains’sensitive to the chemotherapy. This research is aimed at studying the location of theβsubunit of F_1F_0-ATP synthase in tumor cells. At the same time, the effects of the different expression of theβsubunit to cell proliferation has been studied, and the function of F_1F_0-ATP synthase on the surface of tumor cell would be exploited.
In order to study the location of theβsubunit of F_1F_0-ATP synthase,βsubunit was cloned to the vector which contains Fluorescent labels. Extract the total RNA from cells, obtain cDNA by reverse transcript and use PCR to amplify atp5b, which was cloned to the Eukaryotic expression vector pEGFP–N_1. After the transfection, fluorescent was observed on cell surface under the fluorescence microscope.
To investigate the effects of the different expression of theβsubunit to cell proliferation, we designed three shRNA to interfered the expression of ATP5B. Cell was cultured on normal and hypoxic conditions after transfection, cell proliferation was detected by MTT assays. The results show the proliferation rate of cells transfected with shRNA1 or shRNA2 was more than 1, and cells transfected with shRNA3 was less than 1.
引文
[1]张颖娱.ATP合成酶及其功能机制综述[J].分子生物物理学,2004,24 (3):36-40
[2]成君军,黄倢,张士璀.质膜异位F1F0-ATP合成酶研究进展[J].动物医学进展,2008,29(5):60-63
[3]张霞,彭艳,俞丽丽,等.抗人F1F0-ATP合成酶beta亚基单抗的制备及其抗肿瘤活性研究[J].中国免疫学杂志,2008,24:984-992
[4]张吉斌,刘月平,方美英.线粒体ATP合成酶基因组成及生化机制研究进展[J].中国畜牧杂志,2010,46(7):64-68
[5] Wilkens S, Dunn SD, Chandler J, et al. Solution structure of the N-terminal domain of theδsubunit of the E. coli ATP synthase[J]. Nature Struct Biol, 1997, 4: l98- 201
[6] Uhlin U, Cox GB, Guss JM. Crystal structure of theεsubunitof the proton- translocating ATP synthase from Escherichia coli[J]. Structure, 1997, 5: 1219- 1230
[7] Rodgers AJ, Wilkens S, Aggeler R, et al. The subunitδ-subunit b domain of the Escherichia coli F1F0-ATPase. The bsubunits interact with F1 as a dimer and through theδsubunit[J]. J Biol Chem, 1997, 272: 31058-31064
[8] Tatiana VK, Irina GS, Ulf A, et al. Mitochondrial ATP synthase levels in brown adipose tissue are governed by the c-F0 subunit P1 isoform[J]. FASEB J, 2008, 22: 55-63
[9]徐卫红.ATP合成酶及其功能机制综述[J].上饶师范学院学报,2004,24(3):36-40
[10] International Human Genome Sequencing Consortium. Finishing the euchromatic sequence of the human genome[J]. Nature, 2004, 431(7011): 931-945
[11] Griffiths-Jones S, Grocock RJ, van Dongen S, et al. miRBase: micro RNA sequences,targets and gene nomenclature[J]. Nucleic Acids Res, 2006, 34(DATABASE ISSUE): D140- D144
[12] Warburg, O. The Metabolism of Tumors[J]. London: Arnold Constable, 1930, pp: 254–270
[13] Baran AA, Silverman KA, Zeskand J, et al. The modifier of Min 2 (Mom2) locus: Embryonic lethality of a mutation in the Atp5a1 gene suggests a novel mechanism of polyp suppression[J]. Genome Res Co, 2007, 17: 566-576
[14] Hu Dh , Cao K, Wakeman R P, et al. Altered profile of gene expression in rat hearts induced by chronic nicotine consumption[J]. Biochem Bioph Res Co, 297: 729-736
[15] Parikh H, Nilsson E, Ling C, et al. Molecular correlates for maximal oxygen uptake and type1 fibers[J]. Am J Physiol Endocrinol Metab, 2008, 294: E1152-E1159
[16] Ingman M, Kaessmann H, Paabo S, et al. Mitochondrial genome variation and the origin of modern humans[J]. Nature, 2000, 408(6813): 708-713
[17] Jijtmans LG, Henderson NS, Attardi G, et al. Impaired ATP synthase assembly associated with a mutation in the human ATP synthase subunit 6 gene[J]. J Biol Chem, 2001, 276(9): 6755-6762
[18]钱建新.水牛线粒体基因组结构及ATP8基因RNA干扰载体构建[D].广东:中山大学生命科学学院,2006
[19]郝彦哲,杜玉杰,吴夏,等.亚洲黑熊四川亚种(Ursus thibetanus mupinensis)线粒体ATP合成酶和亚基基因克隆及序列分析[J].重庆师范大学学报,2008,25(1):20-23
[20]洪灯,宋绪华,符史干,等.小鼠单个GV期卵母细胞中ATP8基因的表达分析[J].现代生物医学进展,2007,7(10):1488-1490
[21]易平,汪莉,孙清萍,等.红莲型细胞质雄性不育水稻线粒体atp6基因转录本的编辑位点研究[J].生物化学与生物物理进展,2002,29(5):729-733
[22] Zeng XM, Hourset A, Tzagoloff A. The Saccharomyces cerevisiae ATP22 gene codes for the mitochondrial ATPase subunit 6-specific translation factor[J]. Genetics, 2007, 175: 55- 63
[23] Li H S, Zhang J, Thompson BS, et al. Rat mitochondrial ATP synthase ATP5G3: cloning and upregulation in pancreas after chronic ethanol feeding[J]. Physiol Genomics, 2001, 6: 91-98
[24] Zeng XM, Barros MH, Shulman T, et al. ATP25, a new nuclear gene of saccharomyces cerevisiae required for expression and assembly of the atp9p subunit of mitochondrial ATPase[J]. Mol Biol Cell, 2008, 19(4): 1366-1377
[25] Champagne E, Martinez LO, Collet X, et al. Ecto-F1F0 ATP synthase/F1-ATPase: metabolic and immunological functions[J]. Curr Opin Lipidol, 2006, 17(3): 279-284
[26] Das B, Mondragon MO, Sadeghian M, et al. A novel ligand in lymphocyte-mediated cytotoxicity: expression of theβsubunit of H+ transporting ATP synthase on the surface of tumor cell lines[J]. J Exp Med, 1994: 273-281
[27] Moser TL, Kenan DJ, Ashley TA, et al. Endothelial cell surface F1F0-ATP synthase is active in ATP synthesis and is inhibited by angiostatin[J]. Proc Natl Acad Sci, 2001, 8: 6656-6661
[28] Wahl ML, Kenan DJ, Gonzalez-Gronow M, et al. Angiostatin’s molecular mechanism: aspects of specificity and regulation elucidated[J]. J Cell Biochem, 2005, 96(2): 242-261
[29] Kim BW, Choo HJ, Lee JW, et al. Extracellular ATP is generated by ATP synthase complex in adipocyte lipid rafts[J]. Exp Mol Med, 2004, 36(5): 476- 485
[30] Soltys BJ, Kang D, Gupta RS. Localization of P32 protein in mitochondria and at specific ext ramitochondrial locations in normal tissues[J]. Histochem Cell Biol, 2000, 114: 245-255
[31]朱海沫,楼国良.异位F1F0-ATP合成酶及其与肿瘤血管生成的关系[J].中国肿瘤, 2007, 16(8): 610-612
[32]杜艳,高锋.细胞表面ATP合成酶的研究进展[J].上海交通大学学报, 2009, 29, 6: 741-743
[33] Wilson-Fritch L, Burkart A, Bell G, et al. Mitochondrial biogenesis and ermodeling during adipogenesis and in response to the insulin sensitizer rosiglitaazone[J].Mol Cell Biol, 2003, 23(3): 1085-1094
[34] Yu C , Alterman M , Dobrowsky RT. Ceramide displaces cholesterol from lipid rafts and decreasest the association of the cholesterol binding protein caveolin-1[J]. J Lipid Res, 2005, 46: 1678-1691
[35] Wang T, Chen Z, Wang X, et al. Cholesterol loading increases the translocation of ATP synthase beta chain into membrane caveolae in vascular endothelial cells[J]. Biochim Biophys Acta , 2006, 1761: 1182-1190.
[36] Arakaki N, Kita T, Shibata H, et al. Cell-surface H+-ATP synthase as a potential molecular target for anti-obesity durgs[J]. FEBS Lett, 2007, 581(18): 3405-3409
[37] Moser TL, Stack MS, Asplin I,et al. Angiostatin binds ATP synthase on the surface of human endothelial cells[J]. Proc Natl Acad Sci, 1999, 96: 2811-2816
[38] Chi SL, Pizzo SV. Angiostatin is directly cytotoxic to tumor cells at low extracellular pH: a mechanism dependent on cell surface-associated ATP synthase[J]. Cancer Res, 2006, 66: 875-882.
[39] Wahl ML, Grant DS. Effects of microenvironmental extracellular pH and ext- racellular matrix proteins on angiostatin’s activity and on intracellular pH[J]. J Gen Pharm (Vascular ), 2002, 35: 1210
[40] Arakaki N, Nagao T, Niki R, et al. Possible role of cell surface H+-ATP synthase in the extracellular ATP synthesis and proliferation of human umbilical vein endothelial cells ( HUVECs)[J]. Mol Cancer Res, 2003, 1: 931-939
[41] Chi SL, Wahl ML, Mowery YM, et al. Angiostatin-Like activity of a monoclonal antibody to the catalytic subunit of F1F0-ATP synthase[J]. Cancer Res, 2007, 67: 4716-4724
[42] Niina Veitonmaki, Renhai Cao, Lin-Hua Wu, et al. Endothelial Cell Surface ATP Synthase-Triggered Caspase-Apoptotic Pathway Is Essential for K1-5 Induced Antiangiogenesis [J]. Cancer Res , 2004, 64: 3679-3686
[43] Espan A, Berta Mart?n, Ramon Aragues, et al. Bcl-xL Mediated Changes in Metabolic Pathways of Breast Cancer Cells Laura[J]. Am J Pathol, 2005, 167: 1125-1137
[44]李雪飞.大鼠肝卵圆样细胞WB-F344致瘤性转化过程中转录组和蛋白质组的动态变化[D].复旦大学,2008.
[45]孙春晓,于常海.基因功能研究的技术和方法[J].国外医学分子生物学分册,2000,22(2):112-115
[46]卜友泉,杨正梅,宋方洲.新基因功能研究的策略与方法[J].生命科学研究,2006,10(2):95-98
[47]李新枝,蔡佩玲,牟林春.基因功能研究的方法[J].四川解剖学杂志,2007,15(1):57-59
[48]郁兵,淡飞妮,罗雯.RNAi在抗肿瘤研究中的应用[J].科学技术与工程,2009,9(20):6114-6121
[49]王红建,朱金水.RNA干扰技术及其临床应用与进展[J].中国免疫学杂志,2009,25:564-567
[50]朱玉贤,李毅.现代分子生物学[M].高等教育出版社,北京,2002
[51]马元春,张得钧.RNA干扰(RNAi)的研究进展[J].青海医学院学报,2009,30(4):279-282
[52]陈若飞.RNA干扰技术及其应用研究[J].牧与饲料科学,2009,30(5):49-50
[53] Zhan Ma, Manlin Cao, Yiwen Liu, et al. Mitochondrial F1F0-ATP synthase translocates to cell surface in hepatocytes and has high activity in tumor-like acidic and hypoxic environment. Acta Biochim Biophys Sin, 2010, 42: 530–537
[54]马玉珍,石玉涛,白雁,等.EGFP和ALR融合基因表达载体构建及其在HeLa细胞中的表达[J].内蒙古师范大学学报,2006,35(3):340-343
[55]庄振宏,黄缘缘,张峰,等.黄曲霉毒素B1对小鼠肝脏蛋白质组影响的初步研究[J].热带作物学报,2010,31(11):2072-2079
[56]卢绩,陈岐辉,许宁,等.靶向EZH2基因的shRNA真核表达载体的构建及鉴定[J].中国实验诊断学,2010,14(12):1921-1923
[57] Yu JY, Deuiter SL, Turner DL. RNA interference by expression of short interf -ering RNAs and hairpin RNAs in mammalian cells[J]. Proc Natl Acad Sci USA,2002, 99(9): 6047-6052
[58]吴鹏,田媛,桂伶俐,等.RNA干扰技术抑制smad4基因表达对宫颈癌细胞增殖的影响[J].肿瘤,2007,27(3):167-171
[2]成君军,黄倢,张士璀.质膜异位F1F0-ATP合成酶研究进展[J].动物医学进展,2008,29(5):60-63
[3]张霞,彭艳,俞丽丽,等.抗人F1F0-ATP合成酶beta亚基单抗的制备及其抗肿瘤活性研究[J].中国免疫学杂志,2008,24:984-992
[4]张吉斌,刘月平,方美英.线粒体ATP合成酶基因组成及生化机制研究进展[J].中国畜牧杂志,2010,46(7):64-68
[5] Wilkens S, Dunn SD, Chandler J, et al. Solution structure of the N-terminal domain of theδsubunit of the E. coli ATP synthase[J]. Nature Struct Biol, 1997, 4: l98- 201
[6] Uhlin U, Cox GB, Guss JM. Crystal structure of theεsubunitof the proton- translocating ATP synthase from Escherichia coli[J]. Structure, 1997, 5: 1219- 1230
[7] Rodgers AJ, Wilkens S, Aggeler R, et al. The subunitδ-subunit b domain of the Escherichia coli F1F0-ATPase. The bsubunits interact with F1 as a dimer and through theδsubunit[J]. J Biol Chem, 1997, 272: 31058-31064
[8] Tatiana VK, Irina GS, Ulf A, et al. Mitochondrial ATP synthase levels in brown adipose tissue are governed by the c-F0 subunit P1 isoform[J]. FASEB J, 2008, 22: 55-63
[9]徐卫红.ATP合成酶及其功能机制综述[J].上饶师范学院学报,2004,24(3):36-40
[10] International Human Genome Sequencing Consortium. Finishing the euchromatic sequence of the human genome[J]. Nature, 2004, 431(7011): 931-945
[11] Griffiths-Jones S, Grocock RJ, van Dongen S, et al. miRBase: micro RNA sequences,targets and gene nomenclature[J]. Nucleic Acids Res, 2006, 34(DATABASE ISSUE): D140- D144
[12] Warburg, O. The Metabolism of Tumors[J]. London: Arnold Constable, 1930, pp: 254–270
[13] Baran AA, Silverman KA, Zeskand J, et al. The modifier of Min 2 (Mom2) locus: Embryonic lethality of a mutation in the Atp5a1 gene suggests a novel mechanism of polyp suppression[J]. Genome Res Co, 2007, 17: 566-576
[14] Hu Dh , Cao K, Wakeman R P, et al. Altered profile of gene expression in rat hearts induced by chronic nicotine consumption[J]. Biochem Bioph Res Co, 297: 729-736
[15] Parikh H, Nilsson E, Ling C, et al. Molecular correlates for maximal oxygen uptake and type1 fibers[J]. Am J Physiol Endocrinol Metab, 2008, 294: E1152-E1159
[16] Ingman M, Kaessmann H, Paabo S, et al. Mitochondrial genome variation and the origin of modern humans[J]. Nature, 2000, 408(6813): 708-713
[17] Jijtmans LG, Henderson NS, Attardi G, et al. Impaired ATP synthase assembly associated with a mutation in the human ATP synthase subunit 6 gene[J]. J Biol Chem, 2001, 276(9): 6755-6762
[18]钱建新.水牛线粒体基因组结构及ATP8基因RNA干扰载体构建[D].广东:中山大学生命科学学院,2006
[19]郝彦哲,杜玉杰,吴夏,等.亚洲黑熊四川亚种(Ursus thibetanus mupinensis)线粒体ATP合成酶和亚基基因克隆及序列分析[J].重庆师范大学学报,2008,25(1):20-23
[20]洪灯,宋绪华,符史干,等.小鼠单个GV期卵母细胞中ATP8基因的表达分析[J].现代生物医学进展,2007,7(10):1488-1490
[21]易平,汪莉,孙清萍,等.红莲型细胞质雄性不育水稻线粒体atp6基因转录本的编辑位点研究[J].生物化学与生物物理进展,2002,29(5):729-733
[22] Zeng XM, Hourset A, Tzagoloff A. The Saccharomyces cerevisiae ATP22 gene codes for the mitochondrial ATPase subunit 6-specific translation factor[J]. Genetics, 2007, 175: 55- 63
[23] Li H S, Zhang J, Thompson BS, et al. Rat mitochondrial ATP synthase ATP5G3: cloning and upregulation in pancreas after chronic ethanol feeding[J]. Physiol Genomics, 2001, 6: 91-98
[24] Zeng XM, Barros MH, Shulman T, et al. ATP25, a new nuclear gene of saccharomyces cerevisiae required for expression and assembly of the atp9p subunit of mitochondrial ATPase[J]. Mol Biol Cell, 2008, 19(4): 1366-1377
[25] Champagne E, Martinez LO, Collet X, et al. Ecto-F1F0 ATP synthase/F1-ATPase: metabolic and immunological functions[J]. Curr Opin Lipidol, 2006, 17(3): 279-284
[26] Das B, Mondragon MO, Sadeghian M, et al. A novel ligand in lymphocyte-mediated cytotoxicity: expression of theβsubunit of H+ transporting ATP synthase on the surface of tumor cell lines[J]. J Exp Med, 1994: 273-281
[27] Moser TL, Kenan DJ, Ashley TA, et al. Endothelial cell surface F1F0-ATP synthase is active in ATP synthesis and is inhibited by angiostatin[J]. Proc Natl Acad Sci, 2001, 8: 6656-6661
[28] Wahl ML, Kenan DJ, Gonzalez-Gronow M, et al. Angiostatin’s molecular mechanism: aspects of specificity and regulation elucidated[J]. J Cell Biochem, 2005, 96(2): 242-261
[29] Kim BW, Choo HJ, Lee JW, et al. Extracellular ATP is generated by ATP synthase complex in adipocyte lipid rafts[J]. Exp Mol Med, 2004, 36(5): 476- 485
[30] Soltys BJ, Kang D, Gupta RS. Localization of P32 protein in mitochondria and at specific ext ramitochondrial locations in normal tissues[J]. Histochem Cell Biol, 2000, 114: 245-255
[31]朱海沫,楼国良.异位F1F0-ATP合成酶及其与肿瘤血管生成的关系[J].中国肿瘤, 2007, 16(8): 610-612
[32]杜艳,高锋.细胞表面ATP合成酶的研究进展[J].上海交通大学学报, 2009, 29, 6: 741-743
[33] Wilson-Fritch L, Burkart A, Bell G, et al. Mitochondrial biogenesis and ermodeling during adipogenesis and in response to the insulin sensitizer rosiglitaazone[J].Mol Cell Biol, 2003, 23(3): 1085-1094
[34] Yu C , Alterman M , Dobrowsky RT. Ceramide displaces cholesterol from lipid rafts and decreasest the association of the cholesterol binding protein caveolin-1[J]. J Lipid Res, 2005, 46: 1678-1691
[35] Wang T, Chen Z, Wang X, et al. Cholesterol loading increases the translocation of ATP synthase beta chain into membrane caveolae in vascular endothelial cells[J]. Biochim Biophys Acta , 2006, 1761: 1182-1190.
[36] Arakaki N, Kita T, Shibata H, et al. Cell-surface H+-ATP synthase as a potential molecular target for anti-obesity durgs[J]. FEBS Lett, 2007, 581(18): 3405-3409
[37] Moser TL, Stack MS, Asplin I,et al. Angiostatin binds ATP synthase on the surface of human endothelial cells[J]. Proc Natl Acad Sci, 1999, 96: 2811-2816
[38] Chi SL, Pizzo SV. Angiostatin is directly cytotoxic to tumor cells at low extracellular pH: a mechanism dependent on cell surface-associated ATP synthase[J]. Cancer Res, 2006, 66: 875-882.
[39] Wahl ML, Grant DS. Effects of microenvironmental extracellular pH and ext- racellular matrix proteins on angiostatin’s activity and on intracellular pH[J]. J Gen Pharm (Vascular ), 2002, 35: 1210
[40] Arakaki N, Nagao T, Niki R, et al. Possible role of cell surface H+-ATP synthase in the extracellular ATP synthesis and proliferation of human umbilical vein endothelial cells ( HUVECs)[J]. Mol Cancer Res, 2003, 1: 931-939
[41] Chi SL, Wahl ML, Mowery YM, et al. Angiostatin-Like activity of a monoclonal antibody to the catalytic subunit of F1F0-ATP synthase[J]. Cancer Res, 2007, 67: 4716-4724
[42] Niina Veitonmaki, Renhai Cao, Lin-Hua Wu, et al. Endothelial Cell Surface ATP Synthase-Triggered Caspase-Apoptotic Pathway Is Essential for K1-5 Induced Antiangiogenesis [J]. Cancer Res , 2004, 64: 3679-3686
[43] Espan A, Berta Mart?n, Ramon Aragues, et al. Bcl-xL Mediated Changes in Metabolic Pathways of Breast Cancer Cells Laura[J]. Am J Pathol, 2005, 167: 1125-1137
[44]李雪飞.大鼠肝卵圆样细胞WB-F344致瘤性转化过程中转录组和蛋白质组的动态变化[D].复旦大学,2008.
[45]孙春晓,于常海.基因功能研究的技术和方法[J].国外医学分子生物学分册,2000,22(2):112-115
[46]卜友泉,杨正梅,宋方洲.新基因功能研究的策略与方法[J].生命科学研究,2006,10(2):95-98
[47]李新枝,蔡佩玲,牟林春.基因功能研究的方法[J].四川解剖学杂志,2007,15(1):57-59
[48]郁兵,淡飞妮,罗雯.RNAi在抗肿瘤研究中的应用[J].科学技术与工程,2009,9(20):6114-6121
[49]王红建,朱金水.RNA干扰技术及其临床应用与进展[J].中国免疫学杂志,2009,25:564-567
[50]朱玉贤,李毅.现代分子生物学[M].高等教育出版社,北京,2002
[51]马元春,张得钧.RNA干扰(RNAi)的研究进展[J].青海医学院学报,2009,30(4):279-282
[52]陈若飞.RNA干扰技术及其应用研究[J].牧与饲料科学,2009,30(5):49-50
[53] Zhan Ma, Manlin Cao, Yiwen Liu, et al. Mitochondrial F1F0-ATP synthase translocates to cell surface in hepatocytes and has high activity in tumor-like acidic and hypoxic environment. Acta Biochim Biophys Sin, 2010, 42: 530–537
[54]马玉珍,石玉涛,白雁,等.EGFP和ALR融合基因表达载体构建及其在HeLa细胞中的表达[J].内蒙古师范大学学报,2006,35(3):340-343
[55]庄振宏,黄缘缘,张峰,等.黄曲霉毒素B1对小鼠肝脏蛋白质组影响的初步研究[J].热带作物学报,2010,31(11):2072-2079
[56]卢绩,陈岐辉,许宁,等.靶向EZH2基因的shRNA真核表达载体的构建及鉴定[J].中国实验诊断学,2010,14(12):1921-1923
[57] Yu JY, Deuiter SL, Turner DL. RNA interference by expression of short interf -ering RNAs and hairpin RNAs in mammalian cells[J]. Proc Natl Acad Sci USA,2002, 99(9): 6047-6052
[58]吴鹏,田媛,桂伶俐,等.RNA干扰技术抑制smad4基因表达对宫颈癌细胞增殖的影响[J].肿瘤,2007,27(3):167-171