An Investigation into the Potential of Targeting Escherichia coli rne mRNA with Locked Nucleic Acid (LNA) Gapmers as an Antibacterial Strategy.
An Investigation into the Potential of Targeting Escherichia coli rne mRNA with Locked Nucleic Acid (LNA) Gapmers as an Antibacterial Strategy.
复制标题
DOI：
10.3390/molecules26113414
复制
发表时间：
2021-06-04
期刊：
影响因子：
--
通讯作者：
Callaghan AJ
中科院分区：
文献类型：
--
作者：
Goddard LR;Mardle CE;Gneid H;Ball CG;Gowers DM;Atkins HS;Butt LE;Watts JK;Vincent HA;Callaghan AJ
抗菌素耐药性的增加对卫生和国防部门都是一个严峻的挑战，需要新的抗菌靶和抗菌策略。RNA降解和核糖核酸酶，如rne基因编码的必需内切核糖核酸酶E，正在成为潜在的抗菌靶标，而反义寡核苷酸可能提供替代的抗菌策略。由于以前没有使用反义方法来靶向Rne mRNA，我们决定探索使用反义寡核苷酸来靶向大肠杆菌Rne mRNA的翻译起始区。设计合理的反义寡核苷酸，合成锁定核酸(LNA)缺口，通过两种机制抑制Rne基因的翻译。LNA缺口结合可以立体地阻断翻译和/或LNA缺口结合可以促进RNase H介导的Rne mRNA的切割。这可能被证明是一种优势，以前的大多数抗菌反义寡核苷酸方法使用的是将反义寡核苷酸的作用模式限制为空间位阻翻译的寡核苷酸化学方法。利用电泳迁移率改变分析，我们证明了LNA缺口与大肠杆菌Rne mRNA的翻译起始区结合。然后，我们使用无细胞转录翻译报告实验来表明这种结合能够抑制翻译。最后，在体外RNase H切割实验中，LNA裂解物促进了RNase H介导的mRNA切割。尽管反义寡核苷酸传递的挑战仍有待解决，但总的来说，这项工作为开发以反义寡核苷酸靶向Rne mRNA的新的抗菌策略奠定了基础。
The increase in antibacterial resistance is a serious challenge for both the health and defence sectors and there is a need for both novel antibacterial targets and antibacterial strategies. RNA degradation and ribonucleases, such as the essential endoribonuclease RNase E, encoded by the rne gene, are emerging as potential antibacterial targets while antisense oligonucleotides may provide alternative antibacterial strategies. As rne mRNA has not been previously targeted using an antisense approach, we decided to explore using antisense oligonucleotides to target the translation initiation region of the Escherichia coli rne mRNA. Antisense oligonucleotides were rationally designed and were synthesised as locked nucleic acid (LNA) gapmers to enable inhibition of rne mRNA translation through two mechanisms. Either LNA gapmer binding could sterically block translation and/or LNA gapmer binding could facilitate RNase H-mediated cleavage of the rne mRNA. This may prove to be an advantage over the majority of previous antibacterial antisense oligonucleotide approaches which used oligonucleotide chemistries that restrict the mode-of-action of the antisense oligonucleotide to steric blocking of translation. Using an electrophoretic mobility shift assay, we demonstrate that the LNA gapmers bind to the translation initiation region of E. coli rne mRNA. We then use a cell-free transcription translation reporter assay to show that this binding is capable of inhibiting translation. Finally, in an in vitro RNase H cleavage assay, the LNA gapmers facilitate RNase H-mediated mRNA cleavage. Although the challenges of antisense oligonucleotide delivery remain to be addressed, overall, this work lays the foundations for the development of a novel antibacterial strategy targeting rne mRNA with antisense oligonucleotides.
登录
查看更多内容
影响因子：
3.6
作者：
Lee EJ;Groisman EA
通讯作者：
Groisman EA
影响因子：
5.6
作者：
ONO, M;KUWANO, M
通讯作者：
KUWANO, M
影响因子：
1.2
作者：
Kulyté, A;Nekhotiaeva, N;Good, L
通讯作者：
Good, L
影响因子：
5.7
作者：
Lawal A;Jejelowo O;Chopra AK;Rosenzweig JA
通讯作者：
Rosenzweig JA
影响因子：
10.7
作者：
Katoh K;Standley DM
通讯作者：
Standley DM