Garcia-Martin Juan Antonio, Dotu Ivan, Fernandez-Chamorro Javier, Lozano Gloria, Ramajo Jorge, Martinez-Salas Encarnacion, Clote Peter
Biology Department, Boston College, Chestnut Hill, MA 02467, USA.
Department of Experimental and Health Sciences, Research Programme on Biomedical Informatics (GRIB), Universitat Pompeu Fabra. Dr. Aiguader 88, Barcelona, Spain.
Bioinformatics. 2016 Jun 15;32(12):i360-i368. doi: 10.1093/bioinformatics/btw265.
RNA thermometers (RNATs) are cis-regulatory elements that change secondary structure upon temperature shift. Often involved in the regulation of heat shock, cold shock and virulence genes, RNATs constitute an interesting potential resource in synthetic biology, where engineered RNATs could prove to be useful tools in biosensors and conditional gene regulation.
Solving the 2-temperature inverse folding problem is critical for RNAT engineering. Here we introduce RNAiFold2T, the first Constraint Programming (CP) and Large Neighborhood Search (LNS) algorithms to solve this problem. Benchmarking tests of RNAiFold2T against existent programs (adaptive walk and genetic algorithm) inverse folding show that our software generates two orders of magnitude more solutions, thus allowing ample exploration of the space of solutions. Subsequently, solutions can be prioritized by computing various measures, including probability of target structure in the ensemble, melting temperature, etc. Using this strategy, we rationally designed two thermosensor internal ribosome entry site (thermo-IRES) elements, whose normalized cap-independent translation efficiency is approximately 50% greater at 42 °C than 30 °C, when tested in reticulocyte lysates. Translation efficiency is lower than that of the wild-type IRES element, which on the other hand is fully resistant to temperature shift-up. This appears to be the first purely computational design of functional RNA thermoswitches, and certainly the first purely computational design of functional thermo-IRES elements.
RNAiFold2T is publicly available as part of the new release RNAiFold3.0 at https://github.com/clotelab/RNAiFold and http://bioinformatics.bc.edu/clotelab/RNAiFold, which latter has a web server as well. The software is written in C ++ and uses OR-Tools CP search engine.
Supplementary data are available at Bioinformatics online.
RNA温度计（RNATs）是一种顺式调控元件，其二级结构会随温度变化而改变。RNATs常参与热休克、冷休克和毒力基因的调控，在合成生物学中是一种有趣的潜在资源，在合成生物学中，工程化的RNATs可能成为生物传感器和条件基因调控中的有用工具。
解决双温度反向折叠问题对RNAT工程至关重要。在此，我们引入了RNAiFold2T，这是首个用于解决此问题的约束规划（CP）和大邻域搜索（LNS）算法。RNAiFold2T与现有程序（自适应随机游走和遗传算法）反向折叠的基准测试表明，我们的软件生成的解决方案数量多两个数量级，从而能够充分探索解决方案空间。随后，可以通过计算各种指标（包括集合中目标结构的概率、解链温度等）对解决方案进行排序。使用此策略，我们合理设计了两个热传感器内部核糖体进入位点（thermo-IRES）元件，当在网织红细胞裂解物中进行测试时，其标准化的不依赖帽的翻译效率在42°C时比30°C时高约50%。翻译效率低于野生型IRES元件，而野生型IRES元件则完全抗温度升高。这似乎是功能性RNA热开关的首个纯计算设计，当然也是功能性thermo-IRES元件的首个纯计算设计。
RNAiFold2T作为新版本RNAiFold3.0的一部分可在https://github.com/clotelab/RNAiFold和http://bioinformatics.bc.edu/clotelab/RNAiFold上公开获取，后者还有一个网络服务器。该软件用C++编写，并使用OR-Tools CP搜索引擎。
补充数据可在《生物信息学》在线获取。
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