Novel BRD4-NUT fusion isoforms increase the pathogenic complexity in NUT midline carcinoma
Novel BRD4-NUT fusion isoforms increase the pathogenic complexity in NUT midline carcinoma
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DOI：
10.1038/onc.2012.487
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发表时间：
2013-09-26
期刊：
影响因子：
8
通讯作者：
Beesley, A. H.
中科院分区：
文献类型：
--
作者：
Thompson-Wicking, K.;Francis, R. W.;Beesley, A. H.
睾丸核蛋白(NUT)-中线癌(NMC)是一种罕见的侵袭性疾病，典型的表现为单一t(15；19)易位，导致溴结构域包含蛋白4(BRD4)-NUT融合的产生。PER-624是一种来自NMC患者的细胞系，其核型异常复杂，没有给出NUT基因座参与的初步迹象。使用FusionFinder算法对PER-624下一代转录组测序(RNA-Seq)进行分析，发现了一个新的转录本，其中BRD4的外显子15与NUT的外显子2融合，因此与所有已发表的NMC融合转录本不同。PER-624融合中包含的另外三个外显子编码一系列多聚脯氨酸重复序列，其中一个被预测形成螺旋。在NMC细胞系PER-403中，我们鉴定了标准的NMC融合和两种新的异构体。两种细胞系中的小干扰RNA均可抑制细胞增殖，增大细胞大小，表达与上皮细胞分化相一致的细胞角蛋白。这些数据表明，在PER-624中新的BRD4-NUT融合编码了一种功能蛋白，该蛋白是这些细胞中致癌机制的中心。基因组聚合酶链式反应表明，在PER-624和PER-403中，易位都将BRD4的一个内含子融合到NUT编码序列的上游区域。因此，通过易位后RNA剪接产生BRD4-NUT融合转录本似乎是这些癌症的一个共同特征，这一点以前还没有被认识到，其机制促进了不同融合亚型的表达。最后，在PER-403中可以证明野生型NUT的异位表达，这表明了NMC中异常细胞信号的其他途径。这项研究有助于我们了解NMC的遗传多样性，这是为目前治疗难治的疾病找到治疗靶点的重要一步。
Nuclear protein in testis (NUT)-midline carcinoma (NMC) is a rare, aggressive disease typically presenting with a single t(15; 19) translocation that results in the generation of a bromodomain-containing protein 4 (BRD4)-NUT fusion. PER-624 is a cell line generated from an NMC patient with an unusually complex karyotype that gave no initial indication of the involvement of the NUT locus. Analysis of PER-624 next-generation transcriptome sequencing (RNA-Seq) using the algorithm FusionFinder identified a novel transcript in which Exon 15 of BRD4 was fused to Exon 2 of NUT, therefore differing from all published NMC fusion transcripts. The three additional exons contained in the PER-624 fusion encode a series of polyproline repeats, with one predicted to form a helix. In the NMC cell line PER-403, we identified the 'standard' NMC fusion and two novel isoforms. Knockdown by small interfering RNA in either cell line resulted in decreased proliferation, increased cell size and expression of cytokeratins consistent with epithelial differentiation. These data demonstrate that the novel BRD4-NUT fusion in PER-624 encodes a functional protein that is central to the oncogenic mechanism in these cells. Genomic PCR indicated that in both PER-624 and PER-403, the translocation fuses an intron of BRD4 to a region upstream of the NUT coding sequence. Thus, the generation of BRD4-NUT fusion transcripts through post-translocation RNA-splicing appears to be a common feature of these carcinomas that has not previously been appreciated, with the mechanism facilitating the expression of alternative isoforms of the fusion. Finally, ectopic expression of wild-type NUT, a protein normally restricted to the testis, could be demonstrated in PER-403, indicating additional pathways for aberrant cell signaling in NMC. This study contributes to our understanding of the genetic diversity of NMC, an important step towards finding therapeutic targets for a disease that is refractory to current treatments.