Laird Angela S, Mackovski Nikolce, Rinkwitz Silke, Becker Thomas S, Giacomotto Jean
ANZAC Research Institute, Concord Repatriation Hospital, University of Sydney, Sydney, New South Wales, Australia, Department of Biomedical Sciences, Faculty of Medicine & Health Sciences, Macquarie University, Sydney, Australia.
ANZAC Research Institute, Concord Repatriation Hospital, University of Sydney, Sydney, New South Wales, Australia.
Hum Mol Genet. 2016 May 1;25(9):1728-38. doi: 10.1093/hmg/ddw044. Epub 2016 Feb 16.
Spinal muscular atrophy (SMA) is an autosomal recessive disease linked to survival motor neuron (SMN) protein deficiency. While SMN protein is expressed ubiquitously, its deficiency triggers tissue-specific hallmarks, including motor neuron death and muscle atrophy, leading to impaired motor functions and premature death. Here, using stable miR-mediated knockdown technology in zebrafish, we developed the first vertebrate system allowing transgenic spatio-temporal control of the smn1 gene. Using this new model it is now possible to investigate normal and pathogenic SMN function(s) in specific cell types, independently or in synergy with other cell populations. We took advantage of this new system to first test the effect of motor neuron or muscle-specific smn1 silencing. Anti-smn1 miRNA expression in motor neurons, but not in muscles, reproduced SMA hallmarks, including abnormal motor neuron development, poor motor function and premature death. Interestingly, smn1 knockdown in motor neurons also induced severe late-onset phenotypes including scoliosis-like body deformities, weight loss, muscle atrophy and, seen for the first time in zebrafish, reduction in the number of motor neurons, indicating motor neuron degeneration. Taken together, we have developed a new transgenic system allowing spatio-temporal control of smn1 expression in zebrafish, and using this model, we have demonstrated that smn1 silencing in motor neurons alone is sufficient to reproduce SMA hallmarks in zebrafish. It is noteworthy that this research is going beyond SMA as this versatile gene-silencing transgenic system can be used to knockdown any genes of interest, filling the gap in the zebrafish genetic toolbox and opening new avenues to study gene functions in this organism.
脊髓性肌萎缩症（SMA）是一种常染色体隐性疾病，与生存运动神经元（SMN）蛋白缺乏有关。虽然SMN蛋白在全身普遍表达，但其缺乏会引发组织特异性特征，包括运动神经元死亡和肌肉萎缩，导致运动功能受损和过早死亡。在这里，我们利用斑马鱼中稳定的miR介导的敲低技术，开发了首个能对smn1基因进行转基因时空控制的脊椎动物系统。利用这个新模型，现在有可能独立地或与其他细胞群体协同研究特定细胞类型中正常和致病的SMN功能。我们利用这个新系统首先测试了运动神经元或肌肉特异性smn1沉默的效果。在运动神经元而非肌肉中表达抗smn1 miRNA会重现SMA的特征，包括异常的运动神经元发育、运动功能不佳和过早死亡。有趣的是，运动神经元中的smn1敲低还会诱导严重的迟发性表型，包括脊柱侧弯样身体畸形、体重减轻、肌肉萎缩，并且在斑马鱼中首次观察到运动神经元数量减少，表明运动神经元发生了退化。综上所述，我们开发了一种新的转基因系统，可对斑马鱼中smn1的表达进行时空控制，并且利用这个模型，我们证明了仅运动神经元中的smn1沉默就足以在斑马鱼中重现SMA的特征。值得注意的是，这项研究不仅仅局限于SMA，因为这个多功能的基因沉默转基因系统可用于敲低任何感兴趣的基因，填补了斑马鱼遗传工具箱中的空白，并为研究该生物体中的基因功能开辟了新途径。
