小柯机器人

刘如谦团队开发出可用于RNA定点m6A修饰的Cas13工具
2020-06-30 21:19

美国哈佛大学刘如谦(David R. Liu)研究小组开发出可用于RNA定点m6A修饰的Cas13工具。这一研究成果于2020年6月29日在线发表在《自然—生物技术》上。

研究人员发现,具有截短METTL3甲基转移酶结构域的核定位dCas13融合体以及具有修饰功能的METTL3:METTL14甲基转移酶复合物的胞质定位融合体可以在不同细胞区域指导位点特异性m6A的掺入,而前者融合蛋白的脱靶活动特别低。多个位点的独立细胞测定法证实,这种靶向RNA甲基化(TRM)系统以高特异性介导了有效的m6A修饰在内源RNA转录本中。
 
最后,研究人员表明TRM可以诱导m6A介导的转录本丰度变化和选择性剪接。这些发现将TRM确立为用于靶向表观转录组改造的工具,从而可以揭示单个m6A修饰的作用并分析其功能作用。
 
据了解,N6-甲基腺苷(m6A)是人类中最广泛的信使RNA修饰。尽管最近在解析m6A的生物学作用方面取得了进展,但是无法在单个转录本中专门修饰m6A位点,因此仍难以阐明特定m6A的存在与表型结果之间的因果关系。
 
附:英文原文

Title: Programmable m 6 A modification of cellular RNAs with a Cas13-directed methyltransferase

Author: Christopher Wilson, Peter J. Chen, Zhuang Miao, David R. Liu

Issue&Volume: 2020-06-29

Abstract: N6-Methyladenosine (m6A) is the most widespread internal messenger RNA modification in humans. Despite recent progress in understanding the biological roles of m6A, the inability to install m6A site specifically in individual transcripts has hampered efforts to elucidate causal relationships between the presence of a specific m6A and phenotypic outcomes. In the present study, we demonstrate that nucleus-localized dCas13 fusions with a truncated METTL3 methyltransferase domain and cytoplasm-localized fusions with a modified METTL3:METTL14 methyltransferase complex can direct site-specific m6A incorporation in distinct cellular compartments, with the former fusion protein having particularly low off-target activity. Independent cellular assays across multiple sites confirm that this targeted RNA methylation (TRM) system mediates efficient m6A installation in endogenous RNA transcripts with high specificity. Finally, we show that TRM can induce m6A-mediated changes to transcript abundance and alternative splicing. These findings establish TRM as a tool for targeted epitranscriptome engineering that can reveal the effect of individual m6A modifications and dissect their functional roles. The most abundant RNA modification in humans, m6A, can be installed at specified RNA sequences in cells, enabling functional studies.

DOI: 10.1038/s41587-020-0572-6

Source: https://www.nature.com/articles/s41587-020-0572-6

Nature Biotechnology:《自然—生物技术》,创刊于1996年。隶属于施普林格·自然出版集团,最新IF:31.864
官方网址:https://www.nature.com/nbt/
投稿链接:https://mts-nbt.nature.com/cgi-bin/main.plex


本期文章:《自然—生物技术》:Online/在线发表

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