Strand-specific RNA sequencing in Plasmodium falciparum malaria identifies developmentally regulated long non-coding RNA and circular RNA.
Strand-specific RNA sequencing in Plasmodium falciparum malaria identifies developmentally regulated long non-coding RNA and circular RNA.
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DOI：
10.1186/s12864-015-1603-4
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发表时间：
2015-06-13
期刊：
影响因子：
4.4
通讯作者：
Sabeti PC
中科院分区：
文献类型：
--
作者：
Broadbent KM;Broadbent JC;Ribacke U;Wirth D;Rinn JL;Sabeti PC
人类疟疾寄生虫恶性疟原虫具有复杂的多阶段生命周期，需要广泛而精确的基因调控才能入侵和劫持宿主细胞、传播和免疫逃逸。迄今为止，协调这些关键寄生虫过程的监管要素仍然很大程度上未知。然而，越来越清楚的是，长非编码 RNA (lncRNA) 可能代表了广泛生物体中缺失的调控层。为了研究恶性疟原虫中 lncRNA 的调节能力，我们从两个时间过程中收获了 15 个样本。我们的样本集分析了恶性疟原虫 56 小时血液阶段的发育情况。然后，我们开发并验证了链特异性、非 PolyA 选择的 RNA 测序方法，并根据 RNA 测序数据首次组装了恶性疟原虫链特异性转录物结构。这种方法能够注释超过 1000 个 lncRNA 转录本模型及其全面的全局分析：编码预测、周期性、阶段特异性、相关性、GC 含量、长度、相对于注释转录本的位置和剪接。我们验证了三种具有引人注目特性的 lncRNA 的完整剪接结构。非多聚A选择的深度测序还能够预测数百种有趣的恶性疟原虫环状RNA，我们通过实验验证了其中的六种。我们发现，lncRNA 的一个子集，包括所有亚端粒 lncRNA，在侵袭过程中表达达到强烈峰值。相比之下，反义转录物水平在入侵过程中显着下降。与邻近的 mRNA 相比，反义-正义对的表达在血液阶段发育过程中显着反相关，表明转录干扰。我们还验证了恶性疟原虫产生 circRNA，鉴于生物体中缺乏 RNA 干扰，这一点值得注意，并发现高表达的五外显子反义 RNA 有望调节恶性疟原虫配子体发育 1 (PfGDV1)，这是早期性承诺事件所需的基因。本文的在线版本 (doi:10.1186/s12864-015-1603-4) 包含补充材料，可供授权用户使用。
The human malaria parasite Plasmodium falciparum has a complex and multi-stage life cycle that requires extensive and precise gene regulation to allow invasion and hijacking of host cells, transmission, and immune escape. To date, the regulatory elements orchestrating these critical parasite processes remain largely unknown. Yet it is becoming increasingly clear that long non-coding RNAs (lncRNAs) could represent a missing regulatory layer across a broad range of organisms. To investigate the regulatory capacity of lncRNA in P. falciparum, we harvested fifteen samples from two time-courses. Our sample set profiled 56 h of P. falciparum blood stage development. We then developed and validated strand-specific, non-polyA-selected RNA sequencing methods, and pursued the first assembly of P. falciparum strand-specific transcript structures from RNA sequencing data. This approach enabled the annotation of over one thousand lncRNA transcript models and their comprehensive global analysis: coding prediction, periodicity, stage-specificity, correlation, GC content, length, location relative to annotated transcripts, and splicing. We validated the complete splicing structure of three lncRNAs with compelling properties. Non-polyA-selected deep sequencing also enabled the prediction of hundreds of intriguing P. falciparum circular RNAs, six of which we validated experimentally. We found that a subset of lncRNAs, including all subtelomeric lncRNAs, strongly peaked in expression during invasion. By contrast, antisense transcript levels significantly dropped during invasion. As compared to neighboring mRNAs, the expression of antisense-sense pairs was significantly anti-correlated during blood stage development, indicating transcriptional interference. We also validated that P. falciparum produces circRNAs, which is notable given the lack of RNA interference in the organism, and discovered that a highly expressed, five-exon antisense RNA is poised to regulate P. falciparum gametocyte development 1 (PfGDV1), a gene required for early sexual commitment events. The online version of this article (doi:10.1186/s12864-015-1603-4) contains supplementary material, which is available to authorized users.
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