Function of the interacting protein of SMN, the product of the spinal muscular atrophy gene, and neural specific splicing.

SMN蛋白是SMA基因的产物，被认为与SMN相互作用蛋白SIP1在剪接体的生物发生中起着至关重要的作用。为了阐明神经元的剪接机制，我们搜索了神经剪接因子，克隆了两个新的SR蛋白，神经显著SR蛋白(NSSR)1和2，它们在脑和睾丸中表达水平较高。在分化过程中，NSSR-1仅在神经元期表达。纯化的重组NSSR-1和NSSR-2蛋白均能增强核提取液的体外剪接活性。此外，NSSR2的过表达阻止了GluR-B基因的剪接中包含Flip或Flop外显子，导致异常外显子跳过产物的增加。相反，瞬时转染NSSR 1促进了翻转外显子的包含，从而使异常产物被剪接成成熟的剪接形式。结果表明，NSSR1可能在神经元选择性剪接的调控中发挥重要作用。接下来，为了阐明SIP1的发病机制和SIP1的相关性，我们鉴定了SIP1的三个新的剪接变异体(SIP1-β、γ和δ)，以及全长SIP1-α(280AA)。我们检测了这些剪接变异体在各种正常人体组织以及SMA和ALS患者肌肉样本中的表达水平。SIP1-α是一种主要产物，广泛表达。相反，SIP一β和γ的表达水平很低。在肌萎缩侧索硬化症和肌萎缩侧索硬化症患者中，α显著降低(17%，19%)，而β显著升高(34%，32%)。这些发现提示SIP1基因的异常选择性剪接事件在运动神经元疾病中存在，并参与了SMA和ALS的病理过程。

SMN protein, the product of the SMA gene, is considered to play a crucial role in biogenesis of spliceosomes with the SMN-interacting protein, SIP1. To clarify splicing mechanism in neuron, we searched neural splicing factors and cloned two new SR proteins, Neural-salient SR proteins (NSSR) 1 and 2, which are present at higher levels in brain and testis. During the differentiation, NSSR 1 is detected only in the neuronal stage. Both the purified recombinant NSSR 1 and 2 proteins enhance the in vitro splicing activity of nuclear extract. Moreover, overexpression of NSSR 2 prevents the inclusion of either the Flip or Flop exons in the splicing of the GluR-B gene, resulting in an increase in the abnormal exon-skipping product. In contrast, transient transfection with NSSR 1 promotes the inclusion of the Flip exon so that the abnormal product is spliced to the mature spliced form. This suppression of exon skipping by NSSR 1 is observed even with cotransfection of NSSR 2. Results indicate that NSSR 1 may play a crucial role in the regulation of alternative splicing in neurons.Next, to clarify relevancy of pathogenesis and SIP1, we identified three novel splicing variants of the SIP1 (SIP-β、γ and δ), in addition to the full-length SIP1-α (280AA). We examined the expression levels of these splicing variants in various normal human tissues and in muscle samples from patients with SMA and ALS. The SIP1-α was a major product and ubiquitiously expressed. In contrast, SIP-β and γ were detected at very low expression level. In patients with SMA and ALS, the SIP-α was dramatically decreased(17%, 19%) compared to the controls, while the SIP-β was significantly increased(34%, 32%) in both diseases. These findings suggest that aberrant alternative splicing event in SIP1 occur in the motor neuron disease and contribute the pathological process of SMA and ALS.