Parkinson William M, Dookwah Michelle, Dear Mary Lynn, Gatto Cheryl L, Aoki Kazuhiro, Tiemeyer Michael, Broadie Kendal
Department of Biological Sciences, Vanderbilt University, Nashville, TN 37235, USA.
Department of Biochemistry and Molecular Biology, The University of Georgia, Athens, GA 30602, USA.
Dis Model Mech. 2016 May 1;9(5):513-27. doi: 10.1242/dmm.022939. Epub 2016 Mar 3.
Congenital disorders of glycosylation (CDGs) constitute a rapidly growing family of human diseases resulting from heritable mutations in genes driving the production and modification of glycoproteins. The resulting symptomatic hypoglycosylation causes multisystemic defects that include severe neurological impairments, revealing a particularly critical requirement for tightly regulated glycosylation in the nervous system. The most common CDG, CDG-Ia (PMM2-CDG), arises from phosphomannomutase type 2 (PMM2) mutations. Here, we report the generation and characterization of the first Drosophila CDG-Ia model. CRISPR-generated pmm2-null Drosophila mutants display severely disrupted glycosylation and early lethality, whereas RNAi-targeted knockdown of neuronal PMM2 results in a strong shift in the abundance of pauci-mannose glycan, progressive incoordination and later lethality, closely paralleling human CDG-Ia symptoms of shortened lifespan, movement impairments and defective neural development. Analyses of the well-characterized Drosophila neuromuscular junction (NMJ) reveal synaptic glycosylation loss accompanied by defects in both structural architecture and functional neurotransmission. NMJ synaptogenesis is driven by intercellular signals that traverse an extracellular synaptomatrix and are co-regulated by glycosylation and matrix metalloproteinases (MMPs). Specifically, trans-synaptic signaling by the Wnt protein Wingless (Wg) depends on the heparan sulfate proteoglycan (HSPG) co-receptor Dally-like protein (Dlp), which is regulated by synaptic MMP activity. Loss of synaptic MMP2, Wg ligand, Dlp co-receptor and downstream trans-synaptic signaling occurs with PMM2 knockdown. Taken together, this Drosophila CDG disease model provides a new avenue for the dissection of cellular and molecular mechanisms underlying neurological impairments and is a means by which to discover and test novel therapeutic treatment strategies.
糖基化先天性疾病（CDGs）是一类迅速增多的人类疾病，由驱动糖蛋白产生和修饰的基因发生遗传性突变所致。由此产生的症状性低糖基化会导致多系统缺陷，包括严重的神经损伤，这揭示了神经系统中严格调控糖基化的特别关键的需求。最常见的CDG，即CDG-Ia（PMM2-CDG），由磷酸甘露糖变位酶2（PMM2）突变引起。在此，我们报告首个果蝇CDG-Ia模型的构建及特性分析。通过CRISPR技术构建的pmm2基因缺失的果蝇突变体表现出严重紊乱的糖基化和早期致死性，而针对神经元PMM2进行RNA干扰导致寡甘露糖聚糖丰度发生显著变化、渐进性共济失调以及后期致死性，这与人类CDG-Ia的症状（寿命缩短、运动障碍和神经发育缺陷）极为相似。对特征明确的果蝇神经肌肉接头（NMJ）的分析显示，突触糖基化缺失伴随着结构架构和功能性神经传递的缺陷。NMJ突触形成由穿越细胞外突触基质的细胞间信号驱动，并受到糖基化和基质金属蛋白酶（MMPs）的共同调控。具体而言，Wnt蛋白无翅型（Wg）的跨突触信号传导依赖于硫酸乙酰肝素蛋白聚糖（HSPG）共受体类Dally蛋白（Dlp），而Dlp受突触MMP活性的调节。PMM2敲低会导致突触MMP2、Wg配体、Dlp共受体及下游跨突触信号传导缺失。综上所述，这个果蝇CDG疾病模型为剖析神经损伤背后的细胞和分子机制提供了一条新途径，也是发现和测试新型治疗策略的一种手段。
