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Adenovirus VA RNAs impair maturation of primary microRNA

小RNA 小学(天文学) 生物 计算生物学 细胞生物学 遗传学 基因 物理 天文
作者
Ji Hyun Kim,Chang Ho Lee,Seong‐Wook Lee
出处
期刊:Journal of Gene Medicine [Wiley]
卷期号:25 (12)
标识
DOI:10.1002/jgm.3564
摘要

Abstract Background Adenovirus expresses two non‐coding virus‐associated (VA) RNAs: VA I RNA and VA II RNA. Adenovirus‐expressed VA RNAs interfere with the microRNA (miRNA) pathway by competing with precursor miRNAs. The processing pattern of primary miRNA (pri‐miRNA) and factors to affect its processing are not exactly known when using adenovirus for the delivery of pri‐miRNA. Methods To observe pri‐miRNA processing, plasmid construct encoding pri‐miRNA was co‐transfected with VA I/II RNA expression plasmid, or recombinant adenovirus encoding pri‐miRNA was generated and infected. Levels of miRNAs, VA I RNA and VA II RNA were analyzed by a quantitative real‐time PCR (RT‐PCR). VA I–II full‐length RNA was analyzed by a RT‐PCR. RNA immunoprecipitation analysis to pull‐down the VA I–II full‐length RNA binding with Drosha was conducted with Drosha antibody. Results pri‐miRNA was normally processed into mature miRNA when it was expressed in cells using plasmid. However, miRNA maturation was impaired when pri‐miRNA was delivered and expressed using adenovirus. Of note, pri‐miRNA processing was observed to be blocked by VA RNA expression. Such blocked processing could be recovered by introducing antisense RNA of VA RNA, anti‐3′VA RNA. In addition, VA RNAs were transcribed into VA I–II full‐length RNA, which was found to bind and sequester Drosha. Conclusions Adenovirus infection downregulated the processing of pri‐miRNAs in cells, and such downregulation could be derived from VA I–II full‐length RNAs in pri‐miRNA‐like form through competitively binding to Drosha protein. These results indicated that the expression of adenovirus VA RNAs should be inhibited for successful delivery and expression of pri‐miRNA or shRNA in cells using adenovirus.
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