Determinants that regulate splicing of SMN

脊髓性肌萎缩症（SMA）是婴儿死亡的主要遗传原因，但目前还没有治愈方法。 SMA是一种神经肌肉疾病，由运动神经元存活1（SMN 1）缺失引起。近 存在相同的拷贝基因SMN 2，但是，它不能在疾病发展中提供保护。 没有SMN 1。值得注意的是，SMN 1和2编码相同的蛋白质，然而，由于单一的沉默， 突变时，绝大多数SMN 2转录物是可变剪接的，并且最终编码外显子（外显子7： 54 nts）被删除。因此，这种毁灭性疾病的分子基础是一种选择性剪接 导致产生截短且不稳定的SMN蛋白（称为SMN-delta 7）的事件。 通过调节SMN 2剪接模式恢复全长SMN表达代表了一种新的免疫学机制。 治疗干预的令人兴奋的前景。SMA的分子遗传学使这种疾病特别是 适用于促进SMN 2全长表达的治疗策略。1)近99%的 SMA病例由单一基因引起; 2）SMN 2编码相同的蛋白质; 3）SMA人群是 对于SMN 2来说非常均匀。尚未鉴定出纯合子个体 SMN 1和SMN 2为空-可能是因为这种情况将是致命的（与敲- out小鼠模型）。因此，基本上所有SMA患者都携带至少一个SMN 2基因;和4）转录物 SMN 2是稳定的。 该提议的主要目标是开发重组腺相关病毒（rAAV）载体， 表达短RNA，通过促进SMN 2的包含来促进刺激全长SMN表达 外显子7。将使用逐步评估过程来鉴定细胞中最有效的rAAV衍生RNA。 基于模型。最后，将在轻度SMA小鼠模型中评价最有效的rAAV载体，以 确定SMN 2剪接是否可以在体内改变，以及这种增加是否改善了 表征SMA表型。虽然该提案中描述的实验具有直接的 对于SMA治疗的发展的影响，结果可以用作广泛的模型 在这种遗传疾病中，纠正剪接缺陷可以恢复致病基因的功能， 基因

Spinal muscular atrophy (SMA) is the leading genetic cause of infantile death, yet there currently is no cure. SMA is a neuromuscular disorder resulting from the loss of survival motor neuron 1 (SMN1). A nearly identical copy gene exists, SMN2, however, it cannot provide protection from disease development in the absence of SMN1. Remarkably, both SMN1 and 2 encode identical proteins, however, due to a single silent mutation, the vast majority of SMN2 transcripts are alternatively spliced and the final coding exon (exon 7: 54 nts) is removed. Therefore, the molecular basis for this devastating disease is an alternative splicing event that results in the production of a truncated and unstable SMN protein (called SMN-delta7). The restoration of full-length SMN expression by modulating SMN2 splicing patterns represents an exciting prospect for therapeutic intervention. The molecular genetics of SMA make this disease especially amenable to therapeutic strategies that promote full-length expression from SMN2. 1) nearly 99% of all SMA cases are caused by a single gene; 2) SMN2 encodes an identical protein; 3) the SMA population is remarkably homogenous with regards to SMN2. Individuals have not been identified that are homozygous null for SMN1 and SMN2 - presumably because this condition would be lethal (consistent with the knock- out mouse model). Therefore, essentially all SMA patients carry at least one SMN2 gene; and 4) transcripts generated from SMN2 are stable. The primary goal of this proposal is to develop recombinant adeno-associated virus (rAAV) vectors that express short RNAs that promote stimulate full-length SMN expression by promoting the inclusion of SMN2 exon 7. A step-wise evaluation process will be used to identify the most effective rAAV-derived RNAs in cell- based models. Finally, the most effective rAAV vectors will be evaluated in a mild SMA mouse model to determine whether SMN2 splicing can be altered in vivo and whether this increase ameliorates the well- characterized SMA phenotype. While the experiments described in this proposal have immediate implications for the development of a SMA therapy, the results could be used as a model for a broad range of genetic disorders in which correcting a splicing defect would restore functionality to a disease-causing gene.