Translational regulation of Anopheles gambiae mRNAs in the midgut during Plasmodium falciparum infection.
Translational regulation of Anopheles gambiae mRNAs in the midgut during Plasmodium falciparum infection.
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DOI：
10.1186/1471-2164-13-366
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发表时间：
2012-08-02
期刊：
影响因子：
4.4
通讯作者：
Zhu J
中科院分区：
文献类型：
--
作者：
Mead EA;Li M;Tu Z;Zhu J
疟疾是由疟原虫寄生虫引起的，通过受感染的按蚊叮咬传播。中肠入侵是疟原虫在蚊媒体内发育的主要瓶颈。中肠中的疟原虫被富含消化酶的恶劣环境所包围，而快速反应的免疫系统识别疟原虫动合子并从中肠和周围组织招募杀伤因子，从而在它们能够成功穿过中肠上皮之前显著减少入侵动合子的数量。了解寄生虫-载体相互作用的分子细节需要精确测量疟原虫感染期间蚊子中的新生蛋白质合成。目前的表达谱主要监测mRNA稳态水平的变化，但没有解决同样关键的问题，即mRNA编码的蛋白质是否真的合成。在这项研究中，我们使用蔗糖密度梯度离心分离积极翻译冈比亚按蚊mRNA的基础上，他们与多聚核糖体（多聚核糖体）。通过RNA深度测序确定的与多核糖体相关的单个基因转录物的比例反映了mRNA翻译状态。这种方法导致1017蚊子转录本的鉴定，主要是在翻译水平上的恶性疟原虫感染的血液摄入后调节。Caspar是NF-κ B转录因子Rel 2的负调节因子，在疟原虫感染期间似乎在翻译水平上被显著激活。此外，转录Dcr 1，Dcr 2和Drosha，这是参与小RNA的生物合成，表现出增强协会与多核糖体后恶性疟原虫的挑战。这一观察结果表明，蚊子microRNA可能在对抗疟原虫入侵的反应中发挥重要作用。我们分析了总细胞mRNA和与多核糖体相关的mRNA，以同时监测蚊子中的转录组和新生蛋白质合成。这种方法提供了关于蛋白质合成速率的更准确的信息，并确定了一些可能未被识别的蚊子因子，因为这些蛋白质的表达主要在翻译水平上而不是在蚊子摄入疟原虫感染的血餐后的转录水平上受到调节。
Malaria is caused by Plasmodium parasites, which are transmitted via the bites of infected Anopheline mosquitoes. Midgut invasion is a major bottleneck for Plasmodium development inside the mosquito vectors. Malaria parasites in the midgut are surrounded by a hostile environment rich in digestive enzymes, while a rapidly responding immune system recognizes Plasmodium ookinetes and recruits killing factors from the midgut and surrounding tissues, dramatically reducing the population of invading ookinetes before they can successfully traverse the midgut epithelium. Understanding molecular details of the parasite-vector interactions requires precise measurement of nascent protein synthesis in the mosquito during Plasmodium infection. Current expression profiling primarily monitors alterations in steady-state levels of mRNA, but does not address the equally critical issue of whether the proteins encoded by the mRNAs are actually synthesized. In this study, we used sucrose density gradient centrifugation to isolate actively translating Anopheles gambiae mRNAs based upon their association with polyribosomes (polysomes). The proportion of individual gene transcripts associated with polysomes, which is determined by RNA deep sequencing, reflects mRNA translational status. This approach led to identification of 1017 mosquito transcripts that were primarily regulated at the translational level after ingestion of Plasmodium falciparum-infected blood. Caspar, a negative regulator of the NF-kappaB transcription factor Rel2, appears to be substantially activated at the translational levels during Plasmodium infection. In addition, transcripts of Dcr1, Dcr2 and Drosha, which are involved in small RNA biosynthesis, exhibited enhanced associations with polysomes after P. falciparum challenge. This observation suggests that mosquito microRNAs may play an important role in reactions against Plasmodium invasion. We analyzed both total cellular mRNAs and mRNAs that are associated with polysomes to simultaneously monitor transcriptomes and nascent protein synthesis in the mosquito. This approach provides more accurate information regarding the rate of protein synthesis, and identifies some mosquito factors that might have gone unrecognized because expression of these proteins is regulated mainly at the translational level rather than at the transcriptional level after mosquitoes ingest a Plasmodium-infected blood meal.
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