瘤背石磺和里氏拟石磺背部皮肤蛋白质组差异分析
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摘要
从蛋白质水平探讨瘤背石磺和里氏拟石磺背部皮肤蛋白质组差异表达,为完善石磺科贝类从海洋到陆地进化的研究以及解释两种石磺不同的环境适应性提供蛋白质依据。应用Label-free非标定量蛋白质组学技术,结合软件Sequest HT和Proteome Discoverer (Thermo),对瘤背石磺和里氏拟石磺的背部皮肤进行蛋白质组搜库鉴定及定量分析,其中瘤背石磺中鉴定到1 491个蛋白质,里氏拟石磺中鉴定到1 030个蛋白质;用Pfind软件对表达图谱进行同源序列的鉴定及定量分析,检测到928个同源蛋白表达量有差异,瘤背石磺较里氏拟石磺上调表达406个(p<0.05, FC>1.5),下调表达339个(p<0.05, FC<0.5)。适应水下生活的里氏拟石磺背部皮肤角质化程度较低,差异同源蛋白序列分析显示与角质形成、细胞凋亡相关蛋白在里氏拟石磺背部皮肤中高表达,与皮肤保湿性能相关的神经酰胺类物质在瘤背石磺中高表达;KEGG分析显示,嘌呤代谢通路中存在的差异蛋白最多,共73个,其中瘤背石磺较里氏拟石磺上调表达22个,下调表达21个。蛋白组差异分析显示关键酶尿素酶在陆栖性较强的瘤背石磺中高表达,与前人所做动物从水生到陆生进化过程中尿素酶基因完全丢失的研究结论有所出入,属于特殊物种。里氏拟石磺的皮肤辅助呼吸能力和血窦数目均高于瘤背石磺,且背部皮肤中肌纤维更加粗壮,此生理现象在蛋白质组上的表现为两种石磺能量代谢及氧离子运输相关蛋白的差别较大;瘤背石磺背部皮肤中对环境污染反应灵敏的谷胱甘肽S转移酶和谷胱甘肽过氧化物酶的表达量高于里氏拟石磺,推测其与瘤背石磺较强的皮肤免疫能力和环境适应性相关。蛋白表达谱中928个差异表达同源序列为两种石磺表现差异和环境适应的分子机制研究提供了有意义的蛋白质组的基础数据。
Investigating the differential expression of proteome in Onchidium struma and Paraoncidium reevesii dorsal skin at protein level for providing a protein basis by perfecting the study about the evolution of Onchidiidae from sea to wetl and and explaining the different environmental adaptions of two species. Based on Label-free non-standard quantitative proteomic technique and combining with Sequest HT software and Proteome software,the proteome of dorsal skin from O. struma and P. reevesii were identified through the searched database and quantitatively analyzed. 1 491 proteins were identified in O. struma, and 1 030 proteins were identified in P.reevesii. Pfind software was used to identify and quantitatively analyze of homologous sequence in expression map, 928 significant proteins were identified, among which 406 were up regulated(p<0.05, FC>1.5) and 339 were down regulated(p<0.05, FC<0.667) in O. struma compared to P. reevesii. P. reevesii had dorsal skin with lower degree of keratinization level to suit the underwater life, and analysis of homologous protein sequence differences revealed that the proteins related to formation of keratin and apoptosis were highly expressed in dorsal skin of P.reevesii, and that the ceramides associated with moisturizing properties were highly expressed in O. struma. KEGG analysis showed that purine metabolic pathways presented the most differential proteins, total 73 proteins, among which 22 were up-regulated and 21 were down-regulated in O. struma compared to P. reevesii. Proteome differential analysis showed that urease was highly expressed in O. struma, which was mainly terrestrial life. It was different from former research result which urease gene had completely lost in the process of animals from hydrocole to terrestrial evolutionary process. So these two species of Onchidiidae might be special ones. Dorsal skin of P. reevesii had stronger breathing ability, thicker muscle fibers and a greater number of blood sinus than that of O. struma, this physiological phenomenon in proteome on the performance of proteins related to energy metabolism and oxygen ion transport were quite different. The expression of glutathione S transferase and glutathione peroxidase, which was sensitive to the environmental pollution, was higher in dorsal skin of O. struma than that in dorsal skin of P. reevesii, so we speculated that the two proteins bore on O. struma skin immune ability and environmental adaptability. The 928 differentially expressed homologous sequences from protein expression profile might provide significant basic proteome data of two species of Onchidiidae for phenotype differences and environmental adaptation of the molecular mechanism.
Investigating the differential expression of proteome in Onchidium struma and Paraoncidium reevesii dorsal skin at protein level for providing a protein basis by perfecting the study about the evolution of Onchidiidae from sea to wetl and and explaining the different environmental adaptions of two species. Based on Label-free non-standard quantitative proteomic technique and combining with Sequest HT software and Proteome software,the proteome of dorsal skin from O. struma and P. reevesii were identified through the searched database and quantitatively analyzed. 1 491 proteins were identified in O. struma, and 1 030 proteins were identified in P.reevesii. Pfind software was used to identify and quantitatively analyze of homologous sequence in expression map, 928 significant proteins were identified, among which 406 were up regulated(p<0.05, FC>1.5) and 339 were down regulated(p<0.05, FC<0.667) in O. struma compared to P. reevesii. P. reevesii had dorsal skin with lower degree of keratinization level to suit the underwater life, and analysis of homologous protein sequence differences revealed that the proteins related to formation of keratin and apoptosis were highly expressed in dorsal skin of P.reevesii, and that the ceramides associated with moisturizing properties were highly expressed in O. struma. KEGG analysis showed that purine metabolic pathways presented the most differential proteins, total 73 proteins, among which 22 were up-regulated and 21 were down-regulated in O. struma compared to P. reevesii. Proteome differential analysis showed that urease was highly expressed in O. struma, which was mainly terrestrial life. It was different from former research result which urease gene had completely lost in the process of animals from hydrocole to terrestrial evolutionary process. So these two species of Onchidiidae might be special ones. Dorsal skin of P. reevesii had stronger breathing ability, thicker muscle fibers and a greater number of blood sinus than that of O. struma, this physiological phenomenon in proteome on the performance of proteins related to energy metabolism and oxygen ion transport were quite different. The expression of glutathione S transferase and glutathione peroxidase, which was sensitive to the environmental pollution, was higher in dorsal skin of O. struma than that in dorsal skin of P. reevesii, so we speculated that the two proteins bore on O. struma skin immune ability and environmental adaptability. The 928 differentially expressed homologous sequences from protein expression profile might provide significant basic proteome data of two species of Onchidiidae for phenotype differences and environmental adaptation of the molecular mechanism.
引文
Cheng X.Y.,Jiang G.P.,Chai X.L.,and Teng S.S.,2016,Fulllength cDNA cloning and expression analysis of glutathione peroxidase from blood clam Tegillarca granosa,Shuisheng Shengwu Xuebao(Acta Hydrobiologica Sinica),40(6):1144-1151(程雪艳,国萍,柴雪良,滕爽爽,2016,泥蚶谷胱甘肽过氧化物酶基因的全长克隆与表达分析,水生生物学报,40(6):1144-1151)
Dayrat B.,2009,Review of the current knowledge of the systematics of Onchidiidae(Mollusca:Gastropoda:Pulmonata)with a check list of nominal species,Zootaxa,2068:1-26
Fulvio S.,María S.,Marc C.,Gloria G.,Cristina N.,Josep M.,Francesc C.,and Jose M.,2017,Pyruvate kinase M2 and the mitochondrial ATPase inhibitory factor 1 provide novel biomarkers of dermatomyositis:a metabolic link to oncogenesis,Journal of Translational Medicine,15(1):15-29
Ge K.,Lu S.L.,and Qing C.,2005,Arginine metabolism and its effect on woun d healing,Zhongguo Linchuang Yingyang Zazhi(Chinese Journal of Clinical Nutrition),13(1):48-51(葛奎,陆树良,青春,2005,精氨酸代谢及其对创面愈合的影响,中国临床营养杂志,13(1):48-51)
Hammen C.S.,ed.,1980,Marine invertebrates comparative physiology,University Press of New England,London,pp.66-76
Jiang J.J.,Zhang D.C.,Ma J.J.,Su T.F.,and Jiang S.G.,2010,Molecular characterization and expression analysis of a selenium dependent glutathione peroxidase on pearloyster Pinctada fucata,Fujian Nonglin Daxue Xuebao(Journal of Fujian Agriculture and Forestry University(Natural Science E-dition)),39(6):614-621(姜晶晶,张殿昌,麻建军,苏天凤,江世贵,2010,合浦珠母贝含硒谷胱甘肽过氧化物酶基因特征与表达分析,福建农林大学学报(自然科学版),39(6):614-621)
Li K.Q.,and Shao Z.W.,2011,Glycolysis and malignant tumor,Gouji Gukexue Zazhi(Int.J.Orthop.),4(32):243-26(李克乾,邵增务,2011,糖酵解与肿瘤细胞,国际骨科学杂志,4(32):243-26)
Liu C.,Shen H.D.,and Zhou N.,2016,Complete mitochondrial genome of Platevindex sp.(Gastropoda:Pulmonata:Systellommatophora:Onchidiidae),Mitochondrial DNA,27(2):918-920
Liu C.,Wei L.L.,Shen H.D.,Zhu H.C.,and Zhou N.,2015,Complete mitochondrial genome of Peronia verruculata(Gastropoda:Pulmonata:Systellommatophora:Onchidiidae),Mitochondrial DNA,26(5):753-754
Liu R.,2015,The discovery process of glycolysis,Yixue Yu Zhexue(Medicine and Philosoph B),36(18):91-96(刘锐,2015,糖酵解过程的发现史,医学与哲学(B),36(18):91-96)
Liu X.,Shi Y.M.,Wang D.F.,Shen H.D.,Yang T.Z.,Li J.,and Li B.H.,2017,Cloning and tissue expression analysis of the MSTN gene in Onchidium struma,Shuichan Xuebao(Journal of Fisheries of China),41(5):658-668(刘欣,史艳梅,王冬凤,沈和定,杨铁柱,李杰,李柏航,2017,瘤背石磺肌肉生长抑制素基因的克隆及组织表达分析,水产学报,41(5):658-668)
Lv K.L.,Liu D.D.,Zhang H.X.,and Kong X.H.,2015,Function and response to environmental stresses of glutathione peroxidase in aquatic animals,Shuichan Kexue(Fisheries Science),34(6):399-404(吕昆伦,刘丹丹,张红绪,孔祥会,2015,水生动物体内谷胱甘肽过氧化物酶的功能及对环境胁迫的响应,水产科学,34(6):399-404)
Shang P.,2017,Identification of candidate genes on growth traits intergrating transcriptome and proteome of embryonic muscle in pigs,Dissertation for Ph.D.,China Agricultural University,Supervisor:Wu C.X.,pp.39-46(商鹏,2017,基于胚胎肌肉组织转录组和蛋白质组数据鉴定猪生长性状相关基因,博士学位论文,中国农业大学,导师:吴常信,pp.39-46)
Shi F.,and Zhu X.,2012,NOX-mediated MAPK and PI3K/Akt signaling pathways and liver fibrosis,Shijie Huaren Xiaohua Zazhi(World Chinese Journal of Digestology),20(28):2685-2690(施凤,朱萱,2012,NOX介导的MAPK、PI3K/Akt信号通路与肝纤维化的研究进展,世界华人消化杂志,20(28):2685-2690)
Wang D.F.,Shen H.D.,and Wu X.,2015,Comparative histology of the skin from three species of Onchidiidae,Dongwuxue Zazhi(Chinese Journal of Zoology),5(3):437-444(王冬凤,沈和定,吴欣,2015,石磺科3种贝类皮肤显微结构比较,动物学杂志,5(3):437-444)
Yan H.,and Zhou X.Q.,2007,The relationship between the nitrogen metabolism of lysine and aquatic animals,Shuili Yuye(Reservoir Fisheries),27(1):105-107,113(鄢华,周小秋,2007,赖氨酸与水生动物氮代谢的关系,水利渔业,27(1):105-107,113)
Yang Y.,Ma H.J.,and Hu R.,2012,Development of barrier structure and function of stratum corneum,Zhongguo Meirong Yixue(Chinese Journal of Aesthetic Medicine),21(1):158-161(杨扬,马慧军,胡蓉,2012,皮肤角质层的相关屏障结构和功能的研究进展,中国美容医学,21(1):158-161)
Zhang Y.,Cong Y.,and Zhang S.C.,2009,The genes encoding purine metabolism enzymes in amphioxus structure and evolution,Zhongguo Haiyang Daxue Xuebao(Periodical of O-cean University of China),39(1):96-104(张宇,丛艺,张士璀,2009,文昌鱼嘌呤代谢酶基因:结构与演化,中国海洋大学学报(自然科学版),39(1):96-104)
Dayrat B.,2009,Review of the current knowledge of the systematics of Onchidiidae(Mollusca:Gastropoda:Pulmonata)with a check list of nominal species,Zootaxa,2068:1-26
Fulvio S.,María S.,Marc C.,Gloria G.,Cristina N.,Josep M.,Francesc C.,and Jose M.,2017,Pyruvate kinase M2 and the mitochondrial ATPase inhibitory factor 1 provide novel biomarkers of dermatomyositis:a metabolic link to oncogenesis,Journal of Translational Medicine,15(1):15-29
Ge K.,Lu S.L.,and Qing C.,2005,Arginine metabolism and its effect on woun d healing,Zhongguo Linchuang Yingyang Zazhi(Chinese Journal of Clinical Nutrition),13(1):48-51(葛奎,陆树良,青春,2005,精氨酸代谢及其对创面愈合的影响,中国临床营养杂志,13(1):48-51)
Hammen C.S.,ed.,1980,Marine invertebrates comparative physiology,University Press of New England,London,pp.66-76
Jiang J.J.,Zhang D.C.,Ma J.J.,Su T.F.,and Jiang S.G.,2010,Molecular characterization and expression analysis of a selenium dependent glutathione peroxidase on pearloyster Pinctada fucata,Fujian Nonglin Daxue Xuebao(Journal of Fujian Agriculture and Forestry University(Natural Science E-dition)),39(6):614-621(姜晶晶,张殿昌,麻建军,苏天凤,江世贵,2010,合浦珠母贝含硒谷胱甘肽过氧化物酶基因特征与表达分析,福建农林大学学报(自然科学版),39(6):614-621)
Li K.Q.,and Shao Z.W.,2011,Glycolysis and malignant tumor,Gouji Gukexue Zazhi(Int.J.Orthop.),4(32):243-26(李克乾,邵增务,2011,糖酵解与肿瘤细胞,国际骨科学杂志,4(32):243-26)
Liu C.,Shen H.D.,and Zhou N.,2016,Complete mitochondrial genome of Platevindex sp.(Gastropoda:Pulmonata:Systellommatophora:Onchidiidae),Mitochondrial DNA,27(2):918-920
Liu C.,Wei L.L.,Shen H.D.,Zhu H.C.,and Zhou N.,2015,Complete mitochondrial genome of Peronia verruculata(Gastropoda:Pulmonata:Systellommatophora:Onchidiidae),Mitochondrial DNA,26(5):753-754
Liu R.,2015,The discovery process of glycolysis,Yixue Yu Zhexue(Medicine and Philosoph B),36(18):91-96(刘锐,2015,糖酵解过程的发现史,医学与哲学(B),36(18):91-96)
Liu X.,Shi Y.M.,Wang D.F.,Shen H.D.,Yang T.Z.,Li J.,and Li B.H.,2017,Cloning and tissue expression analysis of the MSTN gene in Onchidium struma,Shuichan Xuebao(Journal of Fisheries of China),41(5):658-668(刘欣,史艳梅,王冬凤,沈和定,杨铁柱,李杰,李柏航,2017,瘤背石磺肌肉生长抑制素基因的克隆及组织表达分析,水产学报,41(5):658-668)
Lv K.L.,Liu D.D.,Zhang H.X.,and Kong X.H.,2015,Function and response to environmental stresses of glutathione peroxidase in aquatic animals,Shuichan Kexue(Fisheries Science),34(6):399-404(吕昆伦,刘丹丹,张红绪,孔祥会,2015,水生动物体内谷胱甘肽过氧化物酶的功能及对环境胁迫的响应,水产科学,34(6):399-404)
Shang P.,2017,Identification of candidate genes on growth traits intergrating transcriptome and proteome of embryonic muscle in pigs,Dissertation for Ph.D.,China Agricultural University,Supervisor:Wu C.X.,pp.39-46(商鹏,2017,基于胚胎肌肉组织转录组和蛋白质组数据鉴定猪生长性状相关基因,博士学位论文,中国农业大学,导师:吴常信,pp.39-46)
Shi F.,and Zhu X.,2012,NOX-mediated MAPK and PI3K/Akt signaling pathways and liver fibrosis,Shijie Huaren Xiaohua Zazhi(World Chinese Journal of Digestology),20(28):2685-2690(施凤,朱萱,2012,NOX介导的MAPK、PI3K/Akt信号通路与肝纤维化的研究进展,世界华人消化杂志,20(28):2685-2690)
Wang D.F.,Shen H.D.,and Wu X.,2015,Comparative histology of the skin from three species of Onchidiidae,Dongwuxue Zazhi(Chinese Journal of Zoology),5(3):437-444(王冬凤,沈和定,吴欣,2015,石磺科3种贝类皮肤显微结构比较,动物学杂志,5(3):437-444)
Yan H.,and Zhou X.Q.,2007,The relationship between the nitrogen metabolism of lysine and aquatic animals,Shuili Yuye(Reservoir Fisheries),27(1):105-107,113(鄢华,周小秋,2007,赖氨酸与水生动物氮代谢的关系,水利渔业,27(1):105-107,113)
Yang Y.,Ma H.J.,and Hu R.,2012,Development of barrier structure and function of stratum corneum,Zhongguo Meirong Yixue(Chinese Journal of Aesthetic Medicine),21(1):158-161(杨扬,马慧军,胡蓉,2012,皮肤角质层的相关屏障结构和功能的研究进展,中国美容医学,21(1):158-161)
Zhang Y.,Cong Y.,and Zhang S.C.,2009,The genes encoding purine metabolism enzymes in amphioxus structure and evolution,Zhongguo Haiyang Daxue Xuebao(Periodical of O-cean University of China),39(1):96-104(张宇,丛艺,张士璀,2009,文昌鱼嘌呤代谢酶基因:结构与演化,中国海洋大学学报(自然科学版),39(1):96-104)