生物
甲基转移酶
RNA剪接
核糖核酸
RNA甲基化
细胞生物学
甲基化
RNA结合蛋白
信使核糖核酸
N6-甲基腺苷
融合蛋白
计算生物学
遗传学
基因
重组DNA
作者
Christopher G. Wilson,Peter J. Chen,Miao Zhang,David R. Liu
标识
DOI:10.1038/s41587-020-0572-6
摘要
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.
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