Dual-color STED microscopy reveals a sandwich structure of Bassoon and Piccolo in active zones of adult and aged mice.
Dual-color STED microscopy reveals a sandwich structure of Bassoon and Piccolo in active zones of adult and aged mice.
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双色 STED 显微镜揭示了成年和老年小鼠活动区域中巴松管和短笛的三明治结构。
DOI：
10.1038/srep27935
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发表时间：
2016-06-20
影响因子：
4.6
通讯作者：
Shigemoto K
中科院分区：
文献类型：
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作者：
Nishimune H;Badawi Y;Mori S;Shigemoto K
突触前活动区作为突触传递的突触囊泡释放位点发挥着关键作用，但在亚衍射极限分辨率下哺乳动物神经肌肉接头（NMJ）中活动区的分子结构仍然未知。Bassoon和Piccolo是NMJ、带状突触和脑突触中活性区组装所必需的活性区特异性胞质蛋白。这些蛋白质被认为在活性区共定位并共享一些功能。在这里，我们报告了一个意想不到的发现，这两种蛋白质在小鼠NMJ的非重叠定位显示使用双色受激发射损耗（STED）超分辨率显微镜。短笛点夹在巴松管点之间，并在成年NMJ中排列成短笛-巴松管-短笛结构。P/Q型电压门控钙通道（VGCC）点与大管点共定位。P/Q型VGCC和Bassoon蛋白在老年小鼠NMJ中的表达显著降低。相比之下，来自老年小鼠的NMJ中的Piccolo水平与成年小鼠中的水平相当。本研究在亚衍射极限分辨率下揭示了小鼠NMJs活性区的分子结构，并描述了老年小鼠NMJs活性区蛋白的选择性变性机制。有趣的是，本文所述的活性区蛋白的定位模式类似于使用电子显微镜断层摄影术描述的活性区结构。
Presynaptic active zones play a pivotal role as synaptic vesicle release sites for synaptic transmission, but the molecular architecture of active zones in mammalian neuromuscular junctions (NMJs) at sub-diffraction limited resolution remains unknown. Bassoon and Piccolo are active zone specific cytosolic proteins essential for active zone assembly in NMJs, ribbon synapses, and brain synapses. These proteins are thought to colocalize and share some functions at active zones. Here, we report an unexpected finding of non-overlapping localization of these two proteins in mouse NMJs revealed using dual-color stimulated emission depletion (STED) super resolution microscopy. Piccolo puncta sandwiched Bassoon puncta and aligned in a Piccolo-Bassoon-Piccolo structure in adult NMJs. P/Q-type voltage-gated calcium channel (VGCC) puncta colocalized with Bassoon puncta. The P/Q-type VGCC and Bassoon protein levels decreased significantly in NMJs from aged mouse. In contrast, the Piccolo levels in NMJs from aged mice were comparable to levels in adult mice. This study revealed the molecular architecture of active zones in mouse NMJs at sub-diffraction limited resolution, and described the selective degeneration mechanism of active zone proteins in NMJs from aged mice. Interestingly, the localization pattern of active zone proteins described herein is similar to active zone structures described using electron microscope tomography.