Ago HITS-CLIP expands understanding of Kaposi's sarcoma-associated herpesvirus miRNA function in primary effusion lymphomas.
Ago HITS-CLIP expands understanding of Kaposi's sarcoma-associated herpesvirus miRNA function in primary effusion lymphomas.
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DOI：
10.1371/journal.ppat.1002884
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发表时间：
2012
期刊：
影响因子：
6.7
通讯作者：
Renne R
中科院分区：
文献类型：
--
作者：
Haecker I;Gay LA;Yang Y;Hu J;Morse AM;McIntyre LM;Renne R
KSHV是卡波西肉瘤（KS）、原发性渗出性淋巴瘤（PEL）和多中心Castleman病（MCD）的一个亚类的病原体。KSHV编码的miRNA在所有KSHV相关肿瘤中均易于检测，这一事实表明其在病毒发病机制和肿瘤发生中具有潜在作用。miRNA介导的基因表达调控是一个复杂的网络，每个miRNA都有许多潜在的靶点，迄今为止，只有少数KSHV miRNA靶点已被实验确定。详细了解KSHV miRNA功能需要高通量腐败组学以细胞类型特异性方式全面分析推定的miRNA靶点。我们进行了Ago HITS-CLIP来鉴定两种潜伏性KSHV感染的BEL细胞系中的病毒和细胞miRNA及其同源靶点。Ago HITS-CLIP分别从BCBL-1和BC-3回收了1170和950个细胞KSHVmiRNA靶标。重要的是，富集的簇在许多已充分表征的靶点的3 'UTR中包含KSHV miRNA种子匹配，其中包括THBS 1、BACH 1和C/EBPβ。KSHV miRNA靶点高度富集了参与KSHV生物学中心多个途径的基因，如细胞凋亡、细胞周期调节、淋巴细胞增殖和免疫逃避，从而进一步支持KSHV发病机制和潜在肿瘤发生中的作用。HITS-CLIP还富集了有限数量的病毒转录物，包括仅在潜伏期期间在PEL细胞亚群中表达的vIL-6。有趣的是，Ago HITS-CLIP揭示了极高水平的Ago相关KSHV miRNA，特别是在BC-3细胞中，其中超过70%的所有miRNA是病毒来源的。这表明，除了细胞基因的种子匹配特异性靶向之外，KSHV miRNA还可以通过劫持RISC发挥作用，从而由于人类miRNA的调控丧失而导致细胞基因表达的整体去抑制。总之，我们提供了一个广泛的细胞和病毒miRNA靶点列表，代表了破译KSHV miRNA功能的重要资源。卡波西肉瘤相关疱疹病毒是KS和两种淋巴增生性疾病的病原体：多中心Castleman病和原发性渗出性淋巴瘤（PEL）。KSHV肿瘤是最常见的艾滋病恶性肿瘤，在撒哈拉以南非洲地区，KS是男性中最常见的癌症，无论是否存在HIV感染。KSHV编码12个miRNA基因，其功能在很大程度上未知。病毒miRNAs被整合到RISC中，其主要通过与mRNA的3′ UTR结合来抑制其翻译和/或诱导降解来调节基因表达。迄今为止鉴定的病毒miRNA靶点的小子集表明，这些小的转录后调节因子靶向参与发病机制和肿瘤发生的重要细胞途径。使用Ago HITS-CLIP，一种结合UV交联、Ago-miRNA-mRNA复合物的免疫沉淀和高通量测序的技术，我们在两种常用的研究的PEL细胞系BCBL-1和BC-3中对KSHV miRNA靶组进行了详细分析，并分别鉴定了1170和950个推定的miRNA靶。该数据集提供了一个有价值的资源，以破译KSHV miRNAs如何有助于病毒生物学和发病机制。
KSHV is the etiological agent of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and a subset of multicentricCastleman's disease (MCD). The fact that KSHV-encoded miRNAs are readily detectable in all KSHV-associated tumors suggests a potential role in viral pathogenesis and tumorigenesis. MiRNA-mediated regulation of gene expression is a complex network with each miRNA having many potential targets, and to date only few KSHV miRNA targets have been experimentally determined. A detailed understanding of KSHV miRNA functions requires high-through putribonomics to globally analyze putative miRNA targets in a cell type-specific manner. We performed Ago HITS-CLIP to identify viral and cellular miRNAs and their cognate targets in two latently KSHV-infected PEL cell lines. Ago HITS-CLIP recovered 1170 and 950 cellular KSHVmiRNA targets from BCBL-1 and BC-3, respectively. Importantly, enriched clusters contained KSHV miRNA seed matches in the 3′UTRs of numerous well characterized targets, among them THBS1, BACH1, and C/EBPβ. KSHV miRNA targets were strongly enriched for genes involved in multiple pathways central for KSHV biology, such as apoptosis, cell cycle regulation, lymphocyte proliferation, and immune evasion, thus further supporting a role in KSHV pathogenesis and potentially tumorigenesis. A limited number of viral transcripts were also enriched by HITS-CLIP including vIL-6 expressed only in a subset of PEL cells during latency. Interestingly, Ago HITS-CLIP revealed extremely high levels of Ago-associated KSHV miRNAs especially in BC-3 cells where more than 70% of all miRNAs are of viral origin. This suggests that in addition to seed match-specific targeting of cellular genes, KSHV miRNAs may also function by hijacking RISCs, thereby contributing to a global de-repression of cellular gene expression due to the loss of regulation by human miRNAs. In summary, we provide an extensive list of cellular and viral miRNA targets representing an important resource to decipher KSHV miRNA function. Kaposi's sarcoma-associated herpesvirus is the etiological agent of KS and two lymphoproliferative diseases: multicentricCastleman's disease and primary effusion lymphomas (PEL). KSHV tumors are the most prevalent AIDS malignancies and within Sub-Saharan Africa KS is the most common cancer in males, both in the presence and absence of HIV infection. KSHV encodes 12 miRNA genes whose function is largely unknown. Viral miRNAs are incorporated into RISCs, which regulate gene expression mostly by binding to 3′UTRs of mRNAs to inhibit their translation and/or induce degradation. The small subset of viral miRNA targets identified to date suggests that these small posttranscriptional regulators target important cellular pathways involved in pathogenesis and tumorgenesis. Using Ago HITS-CLIP, a technique which combines UV cross-linking, immunoprecipitation of Ago-miRNA-mRNA complexes, and high throughput sequencing, we performed a detailed analysis of the KSHV miRNA targetome in two commonly studied PEL cell lines, BCBL-1 and BC-3 and identified 1170 and 950 putative miRNA targets, respectively. This data set provides a valuable resource to decipher how KSHV miRNAs contribute to viral biology and pathogenesis.
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影响因子：
64.8
作者：
Baek, Daehyun;Villen, Judit;Shin, Chanseok;Camargo, Fernando D.;Gygi, Steven P.;Bartel, David P.
通讯作者：
Bartel, David P.
影响因子：
13.6
作者：
Di Leva G;Croce CM
通讯作者：
Croce CM
影响因子：
4.8
作者：
An, FQ;Compitello, N;Renne, R
通讯作者：
Renne, R
DOI：
10.1016/j.bbagrm.2011.06.012
发表时间：
2011-11
影响因子：
4.7
作者：
Boss, Isaac W.;Renne, Rolf
通讯作者：
Renne, Rolf
影响因子：
9.2
作者：
Baccarini, Alessia;Chauhan, Hemangini;Gardner, Thomas J.;Jayaprakash, Anitha D.;Sachidanandam, Ravi;Brown, Brian D.
通讯作者：
Brown, Brian D.