Oyaghire Stanley N, Cherubim Collin J, Telmer Cheryl A, Martinez Joe A, Bruchez Marcel P, Armitage Bruce A
Department of Chemistry and Center for Nucleic Acids Science and Technology, Carnegie Mellon University , 4400 Fifth Avenue, Pittsburgh, Pennsylvania 15213-3890, United States.
Biochemistry. 2016 Apr 5;55(13):1977-88. doi: 10.1021/acs.biochem.6b00055. Epub 2016 Mar 18.
We have examined the abilities of three complementary γ-peptide nucleic acid (γPNA) oligomers to invade an RNA G-quadruplex and potently inhibit translation of a luciferase reporter transcript containing the quadruplex-forming sequence (QFS) within its 5'-untranslated region. All three γPNA oligomers bind with low nanomolar affinities to an RNA oligonucleotide containing the QFS. However, while all probes inhibit translation with low to midnanomolar IC50 values, the γPNA designed to hybridize to the first two G-tracts of the QFS and adjacent 5'-overhanging nucleotides was 5-6 times more potent than probes directed to either the 3'-end or internal regions of the target at 37 °C. This position-dependent effect was eliminated after the probes and target were preincubated at an elevated temperature prior to translation, demonstrating that kinetic effects exert significant control over quadruplex invasion and translation inhibition. We also found that antisense γPNAs exhibited similarly potent effects against luciferase reporter transcripts bearing QFS motifs having G2, G3, or G4 tracts. Finally, our results indicate that γPNA oligomers exhibit selectivity and/or potency higher than those of other antisense molecules such as standard PNA and 2'-OMe RNA previously reported to target G-quadruplexes in RNA.
我们研究了三种互补的γ-肽核酸（γPNA）寡聚物侵入RNA G-四链体并有效抑制荧光素酶报告转录本翻译的能力，该转录本在其5'-非翻译区内含有形成四链体的序列（QFS）。所有三种γPNA寡聚物均以低纳摩尔亲和力与含有QFS的RNA寡核苷酸结合。然而，虽然所有探针均以低至中纳摩尔的IC50值抑制翻译，但设计用于与QFS的前两个G-链及相邻的5'-突出核苷酸杂交的γPNA在37°C时比靶向靶标3'-末端或内部区域的探针效力高5-6倍。在翻译前将探针和靶标在升高的温度下预孵育后，这种位置依赖性效应消除了，这表明动力学效应在四链体侵入和翻译抑制中发挥了重要控制作用。我们还发现，反义γPNA对带有具有G2、G3或G4链的QFS基序的荧光素酶报告转录本表现出同样有效的作用。最后，我们的结果表明，γPNA寡聚物表现出比其他反义分子（如先前报道的靶向RNA中G-四链体的标准PNA和2'-OMe RNA）更高的选择性和/或效力。
