Spinal Muscular Atrophy: Is it a motor axon disease?

脊髓性肌萎缩症（SMA）是一种常染色体隐性遗传病，是婴儿死亡的主要原因之一。为了阐明这种运动神经元疾病的生物学机制，在小鼠和斑马鱼身上建立了SMA模型。低水平的存活运动神经元（SMN）蛋白是导致SMA的原因。在斑马鱼中，与低SMN水平相关的最早表型是运动轴突缺陷（截断和异常分支）。这些缺陷是运动轴突的特异性和细胞自主的，并且在运动神经元细胞死亡的情况下发生。这一点，以及其他数据表明，SMA是一种运动轴突疾病。本提案中概述的实验直接验证了这种可能性。首先，将讨论斑马鱼异位运动轴突分支的功能和行为后果。有运动轴突缺陷的个体鱼将被分析为运动缺陷和寿命缩短。为了直接解决运动轴突缺陷是否导致运动神经元细胞死亡，神经肌肉连接、突触活动和运动神经元存活将对单个神经缺陷半节进行分析。其次，通过分析轴突向肋间肌、膈肌和肢体肌的生长情况，研究小鼠SMN耗竭是否会导致运动轴突分支。此外，还将确定在发育过程中何时需要SMN功能来挽救运动轴突缺陷和SMA表型。最后，SMN复合物与RNA转运有关，并存在于生长锥中，这表明SMN可能在将RNA定位到生长锥中以进行局部蛋白质翻译中起作用。在斑马鱼中使用反向遗传方法来消耗运动轴突中与SMN复合物的蛋白质，将揭示减少这些蛋白质是否也会导致运动轴突缺陷和运动神经元细胞死亡。斑马鱼运动轴突也将被用来直接测试当SMN水平降低时，局部蛋白翻译是否会受到影响，使用-肌动蛋白作为候选蛋白。本提案中概述的实验将直接验证SMA是一种运动轴突疾病的假设，并将揭示SMN是否在运动轴突生长锥中对局部蛋白质翻译至关重要的复合体中起作用。

Spinal muscular atrophy (SMA) is an autosomal recessive disease and one of the leading causes of infant mortality. To elucidate the biological mechanism underlying this motoneuron disease, SMA has been modeled in mice and zebrafish. Low levels of the Survival Motor Neuron (SMN) protein are the cause of SMA. The earliest phenotype associated with low SMN levels in zebrafish is motor axon defects (truncations and aberrant branching). These defects are specific and cell-autonomous for motor axons and occur in the absence of motoneuron cell death. This, and data from others showing that SMN is transported down axons to growth cones, indicates that SMA is a motor axon disease. The experiments outlined in this proposal directly test this possibility. First, the functional and behavioral consequences of ectopic motor axon branching in zebrafish will be addressed. Individual fish with defined motor axon defects will be analyzed for movement defects and decreased life span. To address directly whether defective motor axons lead to motoneuronal cell death, neuromuscular junctions, synaptic activity, and motoneuron survival will be analyzed for individual hemisegments with defective nerves. Secondly, whether SMN depletion in mice causes motor axon branching will be examined by analyzing axonal outgrowth into the intercostal, diaphragm, and limb muscles. In addition, it will be determined when in development SMN function is needed to rescue motor axon defects and the SMA phenotype. Lastly, SMN complexes have been implicated in RNA transport and are present in growth cones suggesting that SMN may function in localizing RNAs to growth cones for localized protein translation. Using a reverse genetic approach in zebrafish to deplete proteins shown to complex with SMN in motor axons will reveal whether decreasing these proteins also leads to motor axon defects and motoneuronal cell death. Zebrafish motor axons will also be used to directly test whether localized protein translation is affected when SMN levels are decreased using beta-actin as a candidate protein. Experiments outlined in this proposal will directly test the hypothesis that SMA is a motor axon disease and will reveal whether SMN functions in a complex crucial for localized protein translation in motor axon growth cones.