Diffusion of Large Molecules into Assembling Nuclei Revealed Using an Optical Highlighting Technique
Diffusion of Large Molecules into Assembling Nuclei Revealed Using an Optical Highlighting Technique
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DOI：
10.1016/j.bpj.2009.06.024
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发表时间：
2009-09-02
影响因子：
3.4
通讯作者：
Miyawaki, Atsushi
中科院分区：
文献类型：
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作者：
Shimozono, Satoshi;Tsutsui, Hidekazu;Miyawaki, Atsushi
核膜（NE）定义了核室，NE上的核孔复合物（npc）形成了水通道，水溶性小分子可以被动地扩散。众所周知，小于50 kDa的蛋白质可以通过npc扩散，而大于60 kDa的蛋白质很少通过被动扩散进入。然而，随着NE的重组和核的膨胀，这种大小的界限是如何发展的，我们知之甚少。1987年，一项著名的研究发现了一种有效的机制，通过这种机制，有丝分裂后，大的扩散蛋白（>60 kDa）被排除在重组的细胞核之外。此后，人们普遍认为，在有丝分裂后，新形成的细胞核完全排除了所有蛋白质，除了那些最初与有丝分裂染色体结合的蛋白质和那些通过npc选择性导入的蛋白质。在这里，光转换荧光蛋白KikGR（类似于103 kDa）的四聚体复合体在细胞质中被光学突出显示，并被跟踪以检查其进入细胞核。值得注意的是，突出的复合物在细胞质分裂完成后的20分钟内有效地进入新组装的细胞核。由于KikGR不包含已知的核定位或染色体结合序列，我们的研究结果表明，在核重组过程中，扩散屏障的限制性较小。
The nuclear envelope (NE) defines the nuclear compartment, and nuclear pore complexes (NPCs) on the NE form aqueous passages through which small water-soluble molecules can passively diffuse. It is well known that proteins smaller than 50 kDa can diffuse though NPCs, whereas proteins larger than 60 kDa rarely enter by passive diffusion. Little, however, is known about how this size cutoff develops as the NE reassembles and the nucleus expands. In 1987, a well-known study identified an efficient mechanism by which large diffusing proteins (>60 kDa) were excluded from the reassembling nucleus after mitosis. Since then, it has been generally accepted that after mitosis, newly formed nuclei completely exclude all proteins except those that are initially bound to the mitotic chromosomes and those that are selectively imported through NPCs. Here, the tetrameric complex of the photoconvertible fluorescent protein KikGR (similar to 103 kDa) was optically highlighted in the cytoplasm and followed to examine its entry into nuclei. Remarkably, highlighted complexes efficiently entered newly assembled nuclei during an similar to 20-min period after the completion of cytokinesis. Because KikGR contains no known nuclear-localization or chromosome-binding sequences, our results indicate the diffusion barrier is less restrictive during nuclear reassembly.