Selective Binding to mRNA Duplex Regions by Chemically Modified Peptide Nucleic Acids Stimulates Ribosomal Frameshifting.
Selective Binding to mRNA Duplex Regions by Chemically Modified Peptide Nucleic Acids Stimulates Ribosomal Frameshifting.
复制标题
通过化学修饰的肽核酸选择性结合 mRNA 双链体区域可刺激核糖体移码。
作者：
Ru Ying Puah;Huan Jia;Manikantha Maraswami;Desiree;R. Ero;Lixia Yang;Kiran M. Patil;A. Ong;Manchugondanahalli S. Krishna;Ru;Cailing Tong;Mei Huang;Xin Chen;T. Loh;Yonggui Gao;D. Liu;Gang Chen
负一程序性核糖体移码（-1PRF）允许精确维持病毒蛋白之间的比率，并且参与细胞mRNA半衰期的调节。负一核糖体移码被几个刺激元件激活，如七聚体滑动序列（X XXY YYZ）和位于滑动位点下游2-8个核苷酸的mRNA二级结构（发夹或假结）。在-1 RF时，核糖体阅读框从正常零框移动到-1框，七聚体滑动序列解码为XXX YYY Z而不是X XXY YYZ。我们的研究小组开发了化学修饰的肽核酸（PNA）L和Q单体，通过形成大沟平行PNA·RNA-RNA三链体，分别识别G-C和C-G Watson-Crick碱基对。与单链RNA（ssRNA）相比，掺入L和Q的PNA显示出与双链RNA（dsRNA）的选择性结合。L-和Q-修饰的PNA的序列特异性和结构选择性可以允许精确靶向所需的病毒和细胞RNA结构，因此可以作为有价值的生物学工具用于机制研究和对抗疾病的潜在治疗。在这里，我们首次通过使用兔网织红细胞裂解物的无细胞体外翻译测定证明，靶向模型mRNA发夹的dsRNA特异性化学修饰的PNA刺激-1 RF（从2%至32%）。然而，未修饰的对照PNA显示非特异性的翻译抑制。我们的研究结果表明，修饰的dsRNA结合PNA可能有利于靶向结构化RNA。
Minus-one programmed ribosomal frameshifting (-1 PRF) allows the precise maintenance of the ratio between viral proteins and is involved in the regulation of the half-lives of cellular mRNAs. Minus-one ribosomal frameshifting is activated by several stimulatory elements such as a heptameric slippery sequence (X XXY YYZ) and an mRNA secondary structure (hairpin or pseudoknot) that is positioned 2-8 nucleotides downstream from the slippery site. Upon -1 RF, the ribosomal reading frame is shifted from the normal zero frame to the -1 frame with the heptameric slippery sequence decoded as XXX YYY Z instead of X XXY YYZ. Our research group has developed chemically modified peptide nucleic acid (PNA) L and Q monomers to recognize G-C and C-G Watson-Crick base pairs, respectively, through major-groove parallel PNA·RNA-RNA triplex formation. L- and Q-incorporated PNAs show selective binding to double-stranded RNAs (dsRNAs) over single-stranded RNAs (ssRNAs). The sequence specificity and structural selectivity of L- and Q-modified PNAs may allow the precise targeting of desired viral and cellular RNA structures, and thus may serve as valuable biological tools for mechanistic studies and potential therapeutics for fighting diseases. Here, for the first time, we demonstrate by cell-free in vitro translation assays using rabbit reticulocyte lysate that the dsRNA-specific chemically modified PNAs targeting model mRNA hairpins stimulate -1 RF (from 2% to 32%). An unmodified control PNA, however, shows nonspecific inhibition of translation. Our results suggest that the modified dsRNA-binding PNAs may be advantageous for targeting structured RNAs.