Splicing of a critical exon of human survival motor neuron is regulated by a unique silencer element located in the last intron
Splicing of a critical exon of human survival motor neuron is regulated by a unique silencer element located in the last intron
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DOI：
10.1128/mcb.26.4.1333-1346.2006
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发表时间：
2006-02-01
影响因子：
5.3
通讯作者：
Singh, RN
中科院分区：
文献类型：
--
作者：
Singh, NK;Singh, NN;Singh, RN
人类有两个几乎相同的存活运动神经元(SMN)基因，SMN1和SMN2。在脊髓性肌萎缩症(SMA)中，SMN1不能补偿由于排除外显子7而导致的SMN1的丢失。在这里，我们描述了一个新的位于内含子7的5‘剪接位点下游的抑制元件，我们称之为内含子剪接沉默元件N1(ISS-N1)。ISS-N1的缺失促进了SMN2微型基因来源的mRNAs中的外显子7的包涵体。为了强调ISS-N1在跳过外显子7中的主导作用，当ISS-N1被删除时，允许取消一些积极的顺式元件。证实ISS-N1的沉默功能，ISS-N1是一种针对ISS-N1的反义寡核苷酸，它恢复了SMN2微型基因或内源性SMN2来源的mRNAs中的外显子7包涵体。一直以来，这种寡核苷酸增加了仅携带SMN2基因的SMA患者来源细胞中SMN蛋白的水平。我们的发现首次强调了进化上非保守的内含子元件对SMN2外显子7剪接的深远影响。考虑到内含子序列的寡核苷酸不干扰外显子-连接复合体的形成或mRNA的运输和翻译，ISS-N1为反义寡核苷酸介导的SMN2剪接在SMA中的纠正提供了一个非常特异和有效的治疗靶点。
Humans have two nearly identical copies of the Survival Motor Neuron (SMN) gene, SMN1 and SMN2. In spinal muscular atrophy (SMA), SMN2 is not able to compensate for the loss of SMN1 due to exclusion of exon 7. Here we describe a novel inhibitory element located immediately downstream of the 5' splice site in intron 7. We call this element intronic splicing silencer N1 (ISS-N1). Deletion of ISS-N1 promoted exon 7 inclusion in mRNAs derived from the SMN2 minigene. Underlining the dominant role of ISS-N1 in exon 7 skipping, abrogation of a number of positive cis elements was tolerated when ISS-N1 was deleted. Confirming the silencer function of ISS-N1, an antisense oligonucleotide against ISS-N1 restored exon 7 inclusion in mRNAs derived from the SMN2 minigene or from endogenous SMN2. Consistently, this oligonucleotide increased the levels of SMN protein in SMA patient-derived cells that carry only the SMN2 gene. Our findings underscore for the first time the profound impact of an evolutionarily nonconserved intronic element on SMN2 exon 7 splicing. Considering that oligonucleotides annealing to intronic sequences do not interfere with exon-junction complex formation or mRNA transport and translation, ISS-N1 provides a very specific and efficient therapeutic target for antisense oligonucleotide-mediated correction of SMN2 splicing in SMA.