An RNA targeted to the HIV-1 LTR promoter modulates indiscriminate off-target gene activation.
An RNA targeted to the HIV-1 LTR promoter modulates indiscriminate off-target gene activation.
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DOI：
10.1093/nar/gkm847
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发表时间：
2007
影响因子：
14.9
通讯作者：
Morris KV
中科院分区：
文献类型：
--
作者：
Weinberg MS;Barichievy S;Schaffer L;Han J;Morris KV
转录基因沉默（TGS）可以通过靶向上游启动子区域的小RNA来实现。以前，我们表征了靶向HIV-1长末端重复序列（LTR）启动子247位点的siRNA，并发现siRNA-247（LTR-247 as）的21个碱基的反义链抑制LTR介导的表达。为了表征LTR-247 as的特异性，产生表达靶向跨越247靶位点上游和下游50个碱基的区域的反义RNA的载体。LTR-247 as +7是一种仅在LTR-247 as上游移动7个碱基的反义RNA，其显示了LTR驱动的报告基因表达的显著增加，这与细胞类型和活性染色质甲基标记无关。启动子靶向siRNA最近已显示诱导基因活化。然而，在这里，我们通过序列特异性脱靶效应证明了基因激活。LTR-247 as +7处理的培养物的微阵列分析导致了E185基因的失调。一个功能未知的基因C10 orf 76对LTR-247 as +7的抑制有反应，C10 orf 76的缺失导致几个被LTR-247 as +7激活的基因上调。这些数据表明，当使用短的反义RNA或设计为靶向启动子序列的siRNA时要谨慎，因为启动子靶向RNA可能对具有抑制基因活性的因子具有非预期的抑制作用。
Transcriptional gene silencing (TGS) can be achieved by small RNAs targeted to upstream promoter regions. Previously we characterized siRNAs targeted to the HIV-1 long terminal repeat (LTR) promoter at site 247, and found that a 21-base antisense strand of siRNA-247 (LTR-247as) suppressed LTR-mediated expression. To characterize the specificity of LTR-247as, vectors expressing antisense RNAs targeted to a region spanning 50 bases up- and downstream of the 247 target site were generated. LTR-247as+7, a ∼22 base antisense RNA that is shifted by only seven bases upstream of LTR-247as, showed a significant increase in LTR-driven reporter gene expression that was independent of cell type and active chromatin methyl-marks. Promoter-targeting siRNAs have been recently shown to induce gene activation. However, here we demonstrate gene activation via a sequence-specific off-target effect. Microarray analysis of LTR-247as+7-treated cultures resulted in the deregulation of ∼185 genes. A gene of unknown function, C10orf76, was responsive to inhibition by LTR-247as+7 and the loss of C10orf76 resulted in the upregulation of several genes that were activated by LTR-247as+7. These data suggest caution when using short antisense RNAs or siRNAs designed to target promoter sequences, since promoter-targeted RNAs may have unintended inhibitory effects against factors with suppressive gene activity.
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影响因子：
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作者：
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通讯作者：
Linsley, PS
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