Determinants that regulate splicing of SMN

调节 SMN 剪接的决定因素

• 批准号: 7142158
• 负责人: Christian L. Lorson
• 金额: $ 33.64万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-05-15 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7142158
• 关键词: RNA splicing; animal genetic material tag; binding sites; disease /disorder model; gene expression; gene induction /repression; gene mutation; genetic enhancer element; genetic regulatory element; introns; mutant; nucleic acid sequence; precursor mRNA; progressive spinal muscular atrophy; transcription factor

DESCRIPTION (provided by applicant): 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.

描述（由申请人提供）：脊柱肌肉萎缩（SMA）是婴儿死亡的主要遗传原因，但目前尚无治愈方法。 SMA是由于生存运动神经元1（SMN1）的丧失而导致的神经肌肉疾病。但是，存在一个几乎相同的副本基因SMN2，但是在没有SMN1的情况下，它不能提供免受疾病发展的保护。值得注意的是，SMN1和2均编码相同的蛋白质，但是，由于单个静音突变，绝大多数SMN2转录本被剪接，最终的编码外显子（外显子7：54 NTS）被删除。因此，这种毁灭性疾病的分子基础是替代剪接事件，可导致产生截短且不稳定的SMN蛋白（称为SMN-DELTA7）。通过调节SMN2剪接模式来恢复全长SMN表达，这代表了治疗干预的令人兴奋的前景。 SMA的分子遗传学使该疾病特别适合促进SMN2全长表达的治疗策略。 1）在所有SMA病例中，近99％是由单个基因引起的； 2）SMN2编码相同的蛋白质； 3）SMA人群在SMN2方面非常均匀。尚未鉴定出对SMN1和SMN2的纯合为null的个体 - 大概是因为这种情况是致命的（与敲除小鼠模型一致）。因此，本质上，所有SMA患者至少携带一个SMN2基因。 4）由SMN2产生的转录本稳定。该提案的主要目的是开发重组腺相关病毒（RAAV）向量，表达短RNA，通过促进SMN2 Exon 7的包含来促进刺激全长SMN表达。最后，将在温和的SMA小鼠模型中评估最有效的RAAV矢量，以确定是否可以在体内改变SMN2剪接，以及是否可以改善特征性的SMA表型。尽管该提案中描述的实验对SMA治疗的发展具有直接影响，但结果可以用作广泛遗传疾病的模型，在这种遗传疾病中，纠正剪接缺陷将恢复引起疾病基因的功能。