Simultaneous gene expression profiling in human macrophages infected with Leishmania major parasites using SAGE
Simultaneous gene expression profiling in human macrophages infected with Leishmania major parasites using SAGE
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DOI：
10.1186/1471-2164-9-238
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发表时间：
2008-05-21
期刊：
影响因子：
4.4
通讯作者：
Dellagi, Koussay
中科院分区：
文献类型：
--
作者：
Guerfali, Fatma Z.;Laouini, Dhafer;Dellagi, Koussay
背景：利什曼原虫（L）是细胞内原生动物寄生虫，能够在哺乳动物单核吞噬细胞恶劣且潜在敌意的吞噬溶酶体环境中生存和复制。然后，在努力消除病原体的巨噬细胞 (M Phi) 和为自身生存而挣扎的寄生虫之间发生复杂的相互作用。为了在转录水平上研究这种宿主-寄生虫冲突，在单核细胞来源的人 M Phi s (MDM) 被 L. Major Metacycl 前鞭毛体感染的背景下，采用了基因表达系列分析 (SAGE) 的定量技术 结果：从静息的人类 MF、感染利什曼原虫的人类 MF 和大型利什曼原虫寄生虫中提取 mRNA 后，构建了三个 SAGE 文库并测序，生成多达 28,173 个；分别有 57,514 个和 33,906 个标签（对应于 12,946 个；23,442 个和 9,530 个唯一标签）。通过计算数据分析并与 357,888 个公开的实验人类标签进行直接比较，然后从混合细胞提取物中同时表征寄生虫和宿主细胞转录组，从而自信地区分宿主和寄生虫转录本。该程序使我们能够可靠地将 3,814 个标签分配给 M Phi s 转录本，将 3,666 个标签分配给 L.major 寄生虫转录本。我们重点关注这些，显示其表达的显着变化可能与寄生虫感染的发病机制相关：（i）人类 MF 基因，属于关键免疫反应蛋白（例如 IFN γ 途径、S100 和趋化因子家族）和（ii）一组利什曼原虫基因在寄生虫的细胞内发育阶段优先表达。结论：双重 SAGE 转录组分析 提供了一种有用、强大且准确的方法来区分利什曼原虫感染的人类 M Phi 中人类或寄生虫来源的基因。这项工作中提出的结果表明，利什曼原虫寄生虫调节人类 MF 中的关键转录本，这可能有利于其建立和生存。此外，这些结果提供了寄生虫两个发育阶段（即后循环前鞭毛体和细胞内无鞭毛体）基因表达的概述，并表明它们的转录组谱之间存在广泛差异。最后，我们报告的一组表达基因将在未来几轮数据挖掘和基因注释中有用。
Background: Leishmania (L) are intracellular protozoan parasites that are able to survive and replicate within the harsh and potentially hostile phagolysosomal environment of mammalian mononuclear phagocytes. A complex interplay then takes place between the macrophage (M Phi) striving to eliminate the pathogen and the parasite struggling for its own survival.To investigate this host-parasite conflict at the transcriptional level, in the context of monocyte-derived human M Phi s (MDM) infection by L. major metacyclic promastigotes, the quantitative technique of serial analysis of gene expression (SAGE) was used.Results: After extracting mRNA from resting human MFs, Leishmania-infected human MFs and L. major parasites, three SAGE libraries were constructed and sequenced generating up to 28,173; 57,514 and 33,906 tags respectively (corresponding to 12,946; 23,442 and 9,530 unique tags). Using computational data analysis and direct comparison to 357,888 publicly available experimental human tags, the parasite and the host cell transcriptomes were then simultaneously characterized from the mixed cellular extract, confidently discriminating host from parasite transcripts. This procedure led us to reliably assign 3,814 tags to M Phi s' and 3,666 tags to L. major parasites transcripts. We focused on these, showing significant changes in their expression that are likely to be relevant to the pathogenesis of parasite infection: (i) human MFs genes, belonging to key immune response proteins (e. g., IFN gamma pathway, S100 and chemokine families) and (ii) a group of Leishmania genes showing a preferential expression at the parasite's intra-cellular developing stage.Conclusion: Dual SAGE transcriptome analysis provided a useful, powerful and accurate approach to discriminating genes of human or parasitic origin in Leishmania-infected human M Phi s. The findings presented in this work suggest that the Leishmania parasite modulates key transcripts in human MFs that may be beneficial for its establishment and survival. Furthermore, these results provide an overview of gene expression at two developmental stages of the parasite, namely metacyclic promastigotes and intracellular amastigotes and indicate a broad difference between their transcriptomic profiles. Finally, our reported set of expressed genes will be useful in future rounds of data mining and gene annotation.