Department of Chemistry, Institute of Biochemistry, Westfälische Wilhelms Universität Münster, Corrensstraße 36, 48149, Münster, Germany.
Angew Chem Int Ed Engl. 2023 Jan 26;62(5):e202209975. doi: 10.1002/anie.202209975. Epub 2022 Dec 20.
The 5' cap is a hallmark of eukaryotic mRNA involved in the initiation of translation. Its modification with a single photo-cleavable group can bring translation of mRNA under the control of light. However, UV irradiation causes cell stress and downregulation of translation. Furthermore, complex processes often involve timed expression of more than one gene. The approach would thus greatly benefit from the ability to photo-cleave by blue light and to control more than one mRNA at a time. We report the synthesis of a 5' cap modified with a 7-(diethylamino)coumarin (CouCap) and adapted conditions for in vitro transcription. Translation of the resulting CouCap-mRNA is muted in vitro and in mammalian cells, and can be initiated by irradiation with 450 nm. The native cap is restored and no non-natural residues nor sequence alterations remain in the mRNA. Multiplexing for two different mRNAs was achieved by combining cap analogs with coumarin- and ortho-nitrobenzyl-based photo-cleavable groups.
5' 帽是参与翻译起始的真核 mRNA 的标志。用单个光裂解基团对其进行修饰可以使 mRNA 的翻译受到光的控制。然而，紫外线照射会引起细胞应激和翻译下调。此外，复杂的过程通常涉及同时表达不止一个基因。因此，该方法将极大地受益于能够通过蓝光进行光裂解并同时控制多个 mRNA 的能力。我们报告了用 7-(二乙氨基)香豆素（CouCap）修饰的 5' 帽的合成，并适应了体外转录的条件。所得 CouCap-mRNA 的翻译在体外和哺乳动物细胞中被沉默，并且可以通过 450nm 的照射来启动。天然帽被恢复，mRNA 中没有非天然残基或序列改变。通过将帽类似物与香豆素和邻硝基苄基基光裂解基团结合，实现了两种不同 mRNA 的多重化。
