Identification of LIN28B-bound mRNAs reveals features of target recognition and regulation.
Identification of LIN28B-bound mRNAs reveals features of target recognition and regulation.
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DOI：
10.4161/rna.25194
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发表时间：
2013-07
期刊：
影响因子：
4.1
通讯作者：
Landthaler M
中科院分区：
文献类型：
--
作者：
Graf R;Munschauer M;Mastrobuoni G;Mayr F;Heinemann U;Kempa S;Rajewsky N;Landthaler M
保守的人类 LIN28 RNA 结合蛋白在发育、多能性维持和肿瘤发生中发挥作用。我们使用 PAR-CLIP 和该方法新开发的变体 iDo-PAR-CLIP，以核苷酸分辨率识别人类转录组中 LIN28B 靶标以及由 LIN28B 的各个 RNA 结合域结合的位点。靶标结合位点的位置反映了各个 LIN28B 结合域的已知结构相对方向，从而验证了 iDo-PAR-CLIP。我们的数据表明 LIN28B 直接与大多数表达的 mRNA 和 let-7 microRNA 家族的成员相互作用。 pre-let-7 中检测到的 Lin28 结合基序富含 LIN28B 结合的 mRNA 序列。 LIN28B 敲低后，细胞增殖和细胞周期受到严重损害。 LIN28B 耗尽细胞的定量鸟枪蛋白质组学显示，其 RNA 靶标的蛋白质合成显着减少。计算分析提供的证据表明，蛋白质合成减少的强度与靶转录本中 LIN28B 结合位点的位置相关。
The conserved human LIN28 RNA-binding proteins function in development, maintenance of pluripotency and oncogenesis. We used PAR-CLIP and a newly developed variant of this method, iDo-PAR-CLIP, to identify LIN28B targets as well as sites bound by the individual RNA-binding domains of LIN28B in the human transcriptome at nucleotide resolution. The position of target binding sites reflected the known structural relative orientation of individual LIN28B-binding domains, validating iDo-PAR-CLIP. Our data suggest that LIN28B directly interacts with most expressed mRNAs and members of the let-7 microRNA family. The Lin28-binding motif detected in pre-let-7 was enriched in mRNA sequences bound by LIN28B. Upon LIN28B knockdown, cell proliferation and the cell cycle were strongly impaired. Quantitative shotgun proteomics of LIN28B depleted cells revealed significant reduction of protein synthesis from its RNA targets. Computational analyses provided evidence that the strength of protein synthesis reduction correlated with the location of LIN28B binding sites within target transcripts.