Feed-forward microprocessing and splicing activities at a microRNA-containing intron.
Feed-forward microprocessing and splicing activities at a microRNA-containing intron.
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在含microRNA的内含子处的前馈微孔处理和剪接活性。
DOI：
10.1371/journal.pgen.1002330
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发表时间：
2011-10
期刊：
影响因子：
4.5
通讯作者：
Novina CD
中科院分区：
文献类型：
--
作者：
Janas MM;Khaled M;Schubert S;Bernstein JG;Golan D;Veguilla RA;Fisher DE;Shomron N;Levy C;Novina CD
大多数哺乳动物microRNA（miRNA）基因位于蛋白质编码基因和非编码基因的内含子内，然而协调初级转录物加工成成熟miRNA和剪接mRNA的机制知之甚少。对melastatin内含子6表达的黑色素瘤侵袭抑制因子miR-211的分析显示，miR-211的微加工通过需要Drosha的RNase III活性的机制促进melastatin外显子6-外显子7连接的剪接。此外，内含子6的5′剪接位点（5′SS）的突变，而不是3′SS、分支点或多聚嘧啶段的突变，减少了miR-211的生物合成和Drosha向内含子6的募集，表明剪接体对5′SS的识别促进了miR-211的微加工。在全球范围内，U1剪接因子的敲低降低了内含子miRNA的表达。我们的数据表明，新的相互合作的微加工和剪接活动在内含子的miRNA位点，并表明剪接体组装的启动可能会促进微加工的内含子的miRNA。微RNA（miRNA）基因被转录为含有局部发夹的长的初级RNA，所述发夹被微处理器复合体切除，所述微处理器复合体最少由Drosha和DGCR 8组成。大多数哺乳动物的miRNA位于蛋白质编码和非编码基因的内含子中，但目前还不清楚内含子miRNA的微加工和宿主基因内含子处的剪接如何相互影响。我们最近报道，在黑色素瘤中，melastatin（miR-211）的内含子6表达的miRNA承担其宿主基因的肿瘤抑制功能。在我们目前的工作中，我们检测到melastatin外显子6-外显子7连接相对于其他缺乏内含子miRNAs的外显子-外显子连接升高，表明微加工促进剪接。我们发现，微加工的miR-211之前完成剪接的外显子6-外显子7的连接和Drosha的核酸内切酶的活性，以促进外显子6-外显子7连接的形成。此外，我们发现剪接体组装的第一步，U1 snRNP复合物对5′剪接位点的识别，促进了miR-211和其他内含子而不是基因间miRNA的微加工。我们的研究结果揭示了一个相互合作的，物理的，和功能耦合的内含子miRNA的生物合成和剪接在宿主内含子，他们建议剪接体组装内含子miRNA微加工的整体积极影响。
The majority of mammalian microRNA (miRNA) genes reside within introns of protein-encoding and non-coding genes, yet the mechanisms coordinating primary transcript processing into both mature miRNA and spliced mRNA are poorly understood. Analysis of melanoma invasion suppressor miR-211 expressed from intron 6 of melastatin revealed that microprocessing of miR-211 promotes splicing of the exon 6–exon 7 junction of melastatin by a mechanism requiring the RNase III activity of Drosha. Additionally, mutations in the 5′ splice site (5′SS), but not in the 3′SS, branch point, or polypyrimidine tract of intron 6 reduced miR-211 biogenesis and Drosha recruitment to intron 6, indicating that 5′SS recognition by the spliceosome promotes microprocessing of miR-211. Globally, knockdown of U1 splicing factors reduced intronic miRNA expression. Our data demonstrate novel mutually-cooperative microprocessing and splicing activities at an intronic miRNA locus and suggest that the initiation of spliceosome assembly may promote microprocessing of intronic miRNAs. MicroRNA (miRNA) genes are transcribed as long primary RNAs containing local hairpins that are excised by the Microprocessor complex minimally composed of Drosha and DGCR8. Most mammalian miRNAs reside in introns of protein-encoding and non-coding genes, but it is unclear how microprocessing of an intronic miRNA and splicing at the host gene intron affect each other. We recently reported that in melanoma, a miRNA expressed from intron 6 of melastatin (miR-211) assumes the tumor suppressive function of its host gene. In our current work, we detected elevated melastatin exon 6–exon 7 junctions relative to other exon-exon junctions that lack intronic miRNAs, suggesting that microprocessing promotes splicing. We show that microprocessing of miR-211 precedes completion of splicing of the exon 6–exon 7 junctions and that Drosha's endonuclease activity is required to facilitate exon 6–exon 7 junction formation. Additionally, we found that the first step of spliceosome assembly, recognition of the 5′ splice site by the U1 snRNP complex, promotes microprocessing of miR-211 and other intronic but not intergenic miRNAs. Our findings reveal a mutually cooperative, physical, and functional coupling of intronic miRNA biogenesis and splicing at the host intron, and they suggest a global positive effect of spliceosome assembly on intronic miRNA microprocessing.
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通讯作者：
Novina CD