Multistrand Structure Prediction of Nucleic Acid Assemblies and Design of RNA Switches.
Multistrand Structure Prediction of Nucleic Acid Assemblies and Design of RNA Switches.
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DOI：
10.1021/acs.nanolett.5b04651
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发表时间：
2016-03-09
期刊：
影响因子：
10.8
通讯作者：
Shapiro BA
中科院分区：
文献类型：
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作者：
Bindewald E;Afonin KA;Viard M;Zakrevsky P;Kim T;Shapiro BA
RNA是一种有吸引力的材料，用于创建分子逻辑门，仅在特定分子相互作用伙伴存在的情况下释放编程功能。在这里，我们提出了HyperFold，一种多链RNA/DNA结构预测方法，用于预测可能包含假结的核酸复合物。我们表明，与其他已发表的折叠算法相比，HyperFold的性能也具有竞争力。我们对不同浓度、DNA支点长度和G+C含量进行了大量的RNA/DNA杂交再关联实验，发现观察到的再关联趋势与计算预测相吻合。重要的是，我们将这种方法应用于双链RNA分子开关的设计和实验验证，该开关在与单链RNA支点疾病标志物触发mRNA结合后改变其构象，释放出shrna样的Dicer底物结构。为了证明这一概念，我们分别选择结缔组织生长因子（CTGF） mRNA和增强型绿色荧光蛋白（eGFP） mRNA作为触发序列和靶序列。体外实验证实了RNA开关的形成，并证明当触发RNA与开关支点相互作用时，功能单元被释放。设计的RNA开关被证明在MDA-MB-231乳腺癌细胞中起作用。其他几个开关也被设计和测试。我们得出结论，这种方法具有相当大的潜力，因为原则上，它允许释放针对不同基因的siRNA，这些基因被用作疾病状态的生物标志物。
RNA is an attractive material for the creation of molecular logic gates that release programmed functionalities only in the presence of specific molecular interaction partners. Here we present HyperFold, a multistrand RNA/DNA structure prediction approach for predicting nucleic acid complexes that can contain pseudoknots. We show that HyperFold also performs competitively compared to other published folding algorithms. We performed a large variety of RNA/DNA hybrid reassociation experiments for different concentrations, DNA toehold lengths, and G+C content and find that the observed tendencies for reassociation correspond well to computational predictions. Importantly, we apply this method to the design and experimental verification of a two-stranded RNA molecular switch that upon binding to a single-stranded RNA toehold disease-marker trigger mRNA changes its conformation releasing an shRNA-like Dicer substrate structure. To demonstrate the concept, connective tissue growth factor (CTGF) mRNA and enhanced green fluorescent protein (eGFP) mRNA were chosen as trigger and target sequences, respectively. In vitro experiments confirm the formation of an RNA switch and demonstrate that the functional unit is being released when the trigger RNA interacts with the switch toehold. The designed RNA switch is shown to be functional in MDA-MB-231 breast cancer cells. Several other switches were also designed and tested. We conclude that this approach has considerable potential because, in principle, it allows the release of an siRNA designed against a gene that differs from the gene that is utilized as a biomarker for a disease state.
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DOI：
10.1261/rna.043976.113
发表时间：
2014-06
期刊：
RNA (New York, N.Y.)
影响因子：
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作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
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影响因子：
56.9
作者：
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通讯作者：
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影响因子：
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