王璐;朱顺海;赵其平;黄兵;韩红玉;刘桂玲;李志行;赵焕之;董辉;

• 1:中国农业科学院上海兽医研究所农业农村部动物寄生虫学重点实验室
• 2:上海师范大学生命与环境科学学院

摘要(Abstract)：

柔嫩艾美耳球虫是一种严格的细胞内专性寄生虫,可主动入侵鸡的盲肠上皮细胞。当受到柔嫩艾美耳球虫感染时,宿主细胞会发生相应的变化以应对感染引起的损伤。迄今为止,对宿主细胞应对球虫感染机制的研究极其有限。本研究利用相对和绝对定量同位素标记(iTRAQ)技术结合液相色谱串联质谱(LC-MS/MS)技术分析了感染柔嫩艾美耳球虫子孢子72 h后原代鸡胚成纤维(CEF)细胞的蛋白质组表达变化,结果共鉴定出3967个非冗余蛋白质,与不感染组相比,感染组有259个蛋白质的表达量发生了明显变化,其中上调蛋白质145个和下调蛋白质114个;GO和KEGG通路分析结果发现,这些差异表达蛋白具有结合活性、催化活性、转运子活性等分子功能,主要参与细胞外基质受体互作、糖酵解/糖异生、粘着斑、氨基酸的生物合成、半胱氨酸和蛋氨酸代谢等通路;qPCR结果发现,8个基因的mRNA转录水平变化与iTRAQ结果一致,2个基因的不一致;Western blot结果发现感染组细胞的FABP4蛋白表达量显著降低,与iTRAQ结果中蛋白表达水平变化相一致。本研究结果为柔嫩艾美耳球虫与宿主之间互作关系的深入研究提供了重要的基础数据。

关键词(KeyWords)： iTRAQ;球虫;CEF细胞;差异表达蛋白

Abstract:

Keywords:

基金项目(Foundation): 国家重点研发计划(2017YFD0500400);; 国家寄生虫种质资源共享服务平台(平台-TDRC-22)

作者(Author): 王璐;朱顺海;赵其平;黄兵;韩红玉;刘桂玲;李志行;赵焕之;董辉;

Email:

DOI:

参考文献(References)：

• [1]Shirley M W,Ivens A,Gruber A,et al.The Eimeria genome projects:a sequence of events[J].Trends Parasitol,2004,20(5):199-201.
• [2]Chapman H D.Biochemical,genetic and applied aspects of drug resistance in Eimeria parasites of the fowl[J].Avian Pathol,1997,26(2):221-244.
• [3]Sharman P A,Smith N C,Wallach M G,et al.Chasing the golden egg:vaccination against poultry coccidiosis[J].Parasite Immunol,2010,32(8):590-598.
• [4]Williams R B.A compartmentalised model for the estimation of the cost of coccidiosis to the world's chicken production industry[J].Int J Parasitol,1999,29(8):1209-1229.
• [5]Schmid M,Heitlinger E,Spork S,et al.Eimeria falciformis infection of the mouse caecum identifies opposing roles of IFNγ-regulated host pathways for the parasite development[J].Mucosal Immunol,2014,7(4):969-982.
• [6]Zhang H,Guo F,Zhu G.Involvement of host cell integrin α2 in Cryptosporidium parvum infection[J].Infect Immun,2012,80(5):1753-1758.
• [7]Wang B,Hajano J U,Ren Y,et al.iTRAQ-based quantitative proteomics analysis of rice leaves infected by Rice stripe virus reveals several proteins involved in symptom formation[J].Virol J,2015(12):99.
• [8]朱传智,赵雁林,黄香玉,等.定量蛋白质组学分析链霉素耐药和敏感结核分枝杆菌临床分离株[J].微生物学报,2013,53(2):154-163.
• [9]Witter R L,Solomon J J,Burgoyne G H.Cell culture techniques for primary isolation of Marek's disease-associated herpesvirus[J].Avian Dis,1969,13(1):101-118.
• [10]Wi?niewski J R,Zougman A,Nagaraj N,et al.Universal sample preparation method for proteome analysis[J]Nat Methods,2009,6(5):359-362.
• [11]Zhu M,Dai S,McClung S,et al.Functional differentiation of Brassica napus guard cells and mesophyll cells revealed by comparative proteomics[J].Mol Cell Proteomics,2009,8(4):752-766.
• [12]Ren Y,Hao P,Dutta B,et al.Hypoxia modulates A431 cellular pathways association to tumor radioresistance and enhanced migration revealed by comprehensive proteomic and functional studies[J].Mol Cell Proteomics,2013,12(2):485-498.
• [13]Zhu M,Dai S,McClung S,et al.Functional differentiation of Brassica napus guard cells and mesophyll cells revealed by comparative proteomics[J].Mol Cell Proteomics,2009,8(4):752-766.
• [14]Russell D G,Sinden R E.The role of the cytoskeleton in the motility of coccidian sporozoites[J].J Cell Sci,1981(50):345-359.
• [15]Pradet-Balade B,Boulmé F,Beug H,et al.Translation control:bridging the gap between genomics and proteomics[J]?Trends Biochem Sci,2001,26(4):225-229.
• [16]Wehkamp J,Wang G,Kübler I,et al.The Paneth cell alpha-defensin deficiency of ileal Crohn's disease is linked to Wnt/Tcf-4[J].J Immunol,2007,179(5):3109-3118.
• [17]Sharma S,DeOliveira R B,Kalantari P,et al.Innate immune recognition of an AT-rich stem-loop DNA motif in the Plasmodium falciparum genome[J].Immunity,2011,35(2):194-207.
• [18]Hiscott J,Pitha P,Genin P,et al.Triggering the interferon response:the role of IRF-3 transcription factor[J].J Interferon Cytokine Res,1999,19(1):1-13.
• [19]Honda K,Taniguchi T.IRFs:master regulators of signalling by Toll-like receptors and cytosolic pattern-recognition receptors[J].Nat Rev Immunol,2006,6(9):644-658.
• [20]Laurent F,Lacroix-LamandéS.Innate immune responses play a key role in controlling infection of the intestinal epithelium by Cryptosporidiu[J].Int J Parasitol,2017,47(12):711-721.
• [21]徐慧雯.寄生虫调节宿主免疫应答机制的研究进展[J].世界最新医学信息文摘,2017,17(68):59-61.
• [22]Mangan N E,Fallon R E,Smith P,et al.Helminth infection protects mice from anaphylaxis via IL-10-producing B cells[J].J Immunol,2004,173(10):6346-6356.
• [23]Lépine S,Allegood J C,Edmonds Y,et al.Autophagy induced by deficiency of sphingosine-1-phosphate phosphohydrolase 1 is switched to apoptosis by calpain-mediated autophagy-related gene 5 (Atg5) cleavage[J].J Biol Chem,2011,286(52):44380-44390.
• [24]Yousefi S,Perozzo R,Schmid I,et al.Calpain-mediated cleavage of Atg5 switches autophagy to apoptosis[J].Nat Cell Biol,2006,8(10):1124-1132.
• [25]Zhao Z,Fux B,Goodwin M,et al.Autophagosome-independent essential function for the autophagy protein Atg5 in cellular immunity to intracellular pathogens[J].Cell Host Microbe,2008,4(5):458-469.
• [26]Martin A M,Liu T,Lynn B C,et al.The Toxoplasma gondii parasitophorous vacuole membrane:transactions across the border[J].J Eukaryot Microbiol,2007,54(1):25-28.
• [27]Bultman S,Gebuhr T,Yee D,et al.A Brg1 null mutation in the mouse reveals functional differences among mammalian SWI/SNF complexes[J].Mol Cell,2000,6(6):1287-1295.
• [28]Jacobson R H,Ladurner A G,King D S,et al.Structure and function of a human TAFII250 double bromodomain module[J].Science,2000,288(5470):1422-1425.
• [29]Bakshi R,Hassan M Q,Pratap J,et al.The human SWI/SNF complex associates with RUNX1 to control transcription of hematopoietic target genes[J].J Cell Physiol,2010,225(2):569-576.
• [30]Chen X M,Levine S A,Splinter P L,et al.Cryptosporidium parvum activates nuclear factor κB in biliary epithelia preventing epithelial cell apoptosis[J].Gastroenterology,2001,120(7):1774-1783.
• [31]Heussler V T,Machado J Jr,Fernandez P C,et al.The intracellular parasite Theileria parva protects infected T cells from apoptosis[J].Proc Natl Acad Sci U S A,1999,96(13):7312-7317.
• [32]Lüder C G,Gross U.Apoptosis and its modulation during infection with Toxoplasma gondii:molecular mechanisms and role in pathogenesis[J].Curr Top Microbiol Immunol,2005(289):219-237.
• [33]del Cacho E,Gallego M,López-Bernad F,et al.Expression of anti-apoptotic factors in cells parasitized by second-generation schizonts of Eimeria tenella and Eimeria necatrix[J].Vet Parasitol,2004,125(3-4):287-300.