Transcriptomic and proteomic analyses of rhabdomyosarcoma cells reveal differential cellular gene expression in response to enterovirus 71 infection
Transcriptomic and proteomic analyses of rhabdomyosarcoma cells reveal differential cellular gene expression in response to enterovirus 71 infection
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DOI：
10.1111/j.1462-5822.2005.00644.x
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发表时间：
2006-04-01
影响因子：
3.4
通讯作者：
Chow, VTK
中科院分区：
文献类型：
--
作者：
Leong, WF;Chow, VTK
通过阐明宿主和病毒介导的转录反应，可以深入了解宿主抗病毒策略以及病毒疾病表现。采用基于寡核苷酸的微阵列分析感染肠道病毒 71 (EV71) MS/7423/87 株的横纹肌肉瘤细胞在感染后 20 小时的 mRNA。使用 Acuity 软件和 LOWESS 标准化，发现 152 个基因下调，而 39 个基因上调两倍以上。改变的转录本包括编码细胞骨架成分、蛋白质翻译和修饰的转录本；细胞转运蛋白；蛋白质降解介质；细胞死亡介质；线粒体相关蛋白和代谢蛋白；细胞受体和信号传感器。通过实时逆转录聚合酶链式反应 (RT-PCR) 验证了 15 个代表性基因表达谱的变化，该反应还比较了感染 EV71 毒株 5865/Sin/000009 的细胞的转录反应，该毒株是从 2000 年新加坡爆发期间的致命病例中分离出来的。APOB、CLU、DCAMKL1 和 ODC1 蛋白的蛋白质印迹分析与蛋白质和转录水平相关。二维蛋白质组图突出显示了 EV71 感染后细胞蛋白（CCT5、CFL1、ENO1、HSPB1、PSMA2 和 STMN1）表达的差异。一些细胞凋亡相关基因的表达发生了改变，与脊髓灰质炎病毒感染中观察到的细胞凋亡减弱相一致。有趣的是，参与大脑发育的双皮质素和 CaM 激酶样 1 (DCAMKL1) 在感染过程中高度表达。因此，微阵列、实时 RT-PCR 和蛋白质组分析可以阐明导致 EV71 发病机制的众多复杂细胞反应的整体观点。
Insights into the host antiviral strategies as well as viral disease manifestations can be achieved through the elucidation of host- and virus-mediated transcriptional responses. An oligo-based microarray was employed to analyse mRNAs from rhabdomyosarcoma cells infected with the MS/7423/87 strain of enterovirus 71 (EV71) at 20 h post infection. Using Acuity software and LOWESS normalization, 152 genes were found to be downregulated while 39 were upregulated by greater than twofold. Altered transcripts include those encoding components of cytoskeleton, protein translation and modification; cellular transport proteins; protein degradation mediators; cell death mediators; mitochondrial-related and metabolism proteins; cellular receptors and signal transducers. Changes in expression profiles of 15 representative genes were authenticated by real-time reverse transcription polymerase chain reaction (RT-PCR), which also compared the transcriptional responses of cells infected with EV71 strain 5865/Sin/000009 isolated from a fatal case during the Singapore outbreak in 2000. Western blot analyses of APOB, CLU, DCAMKL1 and ODC1 proteins correlated protein and transcript levels. Two-dimensional proteomic maps highlighted differences in expression of cellular proteins (CCT5, CFL1, ENO1, HSPB1, PSMA2 and STMN1) following EV71 infection. Expression of several apoptosis-associated genes was modified, coinciding with apoptosis attenuation observed in poliovirus infection. Interestingly, doublecortin and CaM kinase-like 1 (DCAMKL1) involved in brain development, was highly expressed during infection. Thus, microarray, real-time RT-PCR and proteomic analyses can elucidate the global view of the numerous and complex cellular responses that contribute towards EV71 pathogenesis.