支原体
细胞内
寡核苷酸
基因表达
核糖核酸
分枝杆菌
抄写(语言学)
基因
反义RNA
分子生物学
生物
化学
生物化学
反义治疗
结核分枝杆菌
微生物学
细菌
肺结核
医学
病理
锁核酸
哲学
遗传学
语言学
作者
Yulia V. Skvortsova,Elena G. Salina,Е. А. Буракова,Oksana S. Bychenko,Dmitry A. Stetsenko,Tatyana Azhikina
标识
DOI:10.3389/fphar.2019.01049
摘要
The worldwide spread of multidrug-resistant Mycobacterium tuberculosis strains prompted the development of new strategies to combat tuberculosis, one of which is antisense therapy based on targeting bacterial mRNA by oligonucleotide derivatives. However, the main limitation of antisense antibacterials is poor cellular uptake because of electrostatic charge. Phosphoryl guanidine oligo-2'-O-methylribonucleotides (2'-OMe PGOs) are a novel type of uncharged RNA analogues with high RNA affinity, which penetrate through the bacterial cell wall more efficiently. In this study, we investigated the uptake and biological effects of 2'-OMe PGO in mycobacteria. The results indicated that 2'-OMe PGO specific for the alanine dehydrogenase-encoding ald gene inhibited the growth of Mycobacterium smegmatis and downregulated ald expression at both the transcriptional and translational levels through an RNase H-independent mechanism, showing higher biological activity than its phosphorothioate oligonucleotide counterpart. Confocal microscopy revealed that the anti-ald 2'-OMe PGO was taken up by intracellular mycobacteria residing in RAW 264.7 macrophages without exerting toxic effects on eukaryotic cells, indicating that 2'-OMe PGO was able to efficiently cross two cellular membranes. In addition, 2'-OMe PGO inhibited the transcription of the target ald gene in M. smegmatis-infected macrophages. Thus, we demonstrated, for the first time, a possibility of targeting gene expression and inhibiting growth of intracellular mycobacteria by antisense oligonucleotide derivatives. Strong antisense activity and efficient uptake of the new RNA analogue, 2'-OMe PGO, by intracellular microorganisms revealed here may promote the development of novel therapeutic strategies to treat TB and prevent the emergence of drug-resistant mycobacterial strains.
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