Development of a method for regulating transcription by light sensitive hairpin-type peptide nucleic acid

我们合成了一种发夹型多肽核酸-AZO(发夹型PNA-AZO)，它用可见光敏的偶氮苯连接物连接Watson-Crick链和Hoogsteen链。我们证实偶氮苯连接体增强了与其互补DNA序列的亲和力，并与其目标DNA序列形成了1：1的复合体。发夹型PNA-AZO的可见光照射改变了其与靶DNA的结合性质，与顺式异构体相比，反式异构体表现出更高的结合亲和力。我们进一步用荧光显微镜检测了发夹型PNA-AZO对H1N1流感病毒血凝素(HA)基因和维多利亚Aequorea基因绿色荧光蛋白(GFP)基因在HeLa细胞中转录的调控。结果表明，发夹型PNA-AZO反式表达降低了细胞内HA-GFP蛋白的表达，而顺式PNA-AZO不干扰HA-GFP蛋白的表达。根据这些结果，我们可以通过可见光照射控制发夹型PNA-AZO的偶氮苯单元的构型来调节DNA在细胞中的转录。

We synthesized a hairpin-type peptide nucleic acid-AZO(hairpin-type PNA-AZO) that tethers a Watson-Crick strand and a Hoogsteen strand with visible light-sensitive azobenzene linkers. We confirmed the azobenzene linkers enhanced the affinity to its complementary DNA sequence and afforded a 1;1 complex formation with its target DNA sequence. Visible light irradiation to the hairpin-type PNA-AZO altered its binding property to the target DNA and thetrans-isoform exhibited higher binding affinity compared to the cis-isoform. We further examined if hairpin-type-PNA-AZO can regulate transcription of plasmid encoding hemagglutinine(HA) protein of influenza A/H1N1 virus and green fluorescence protein(GFP) of Aequorea victoria in HeLa cell by fluorescence microscopy. As a result, the trans-form of hairpin-type PNA-AZO reduced the expression of HA-GFP protein in cells, while the cis-form didn’t interfer the expression. From these results, we can regulatetranscription of DNA in cells by controlling the configuration of azobenzene unit of hairpin-type PNA-AZO via visible light irradiation.