TRANSLOCATION OF RNA-COATED GOLD PARTICLES THROUGH THE NUCLEAR-PORES OF OOCYTES
TRANSLOCATION OF RNA-COATED GOLD PARTICLES THROUGH THE NUCLEAR-PORES OF OOCYTES
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DOI：
10.1083/jcb.106.3.575
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发表时间：
1988-03-01
影响因子：
7.8
通讯作者：
FELDHERR, CM
中科院分区：
文献类型：
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作者：
DWORETZKY, SI;FELDHERR, CM
在本研究中，不同大小的包被tRNA、5S RNA或聚(A)的金颗粒被用来定位和表征RNA转移到细胞质的途径。将包被RNA的金颗粒显微注射到非洲爪哇卵母细胞的核中。在15、60min或6h后固定细胞，然后用电子显微镜观察颗粒分布。所有类别的RNA都得到了类似的结果。核注射后，在核孔中央通道内和紧邻核孔的细胞质内可观察到直径20-230微米的颗粒。这种大小的颗粒不会通过毛孔扩散，这表明发生了某种形式的中介运输。此外，还发现易位过程是可饱和的。至少97%的被分析毛孔似乎参与了易位过程。包被非生理性多核苷酸(聚[I]或聚[达])的金也发生了移位。当细胞核被注入牛血清白蛋白、卵清蛋白、聚谷氨酸或PVP包被的金时，这些颗粒基本上被排除在毛孔之外。这些结果表明，RNA-金在气孔和邻近细胞质中的积累不是由于非特异性效应。我们的结论是，包被不同类别RNA的金颗粒的易位位置位于核孔的中心，直径至少230°的颗粒可以穿过包膜。在注射部位附近区域的核孔内可观察到注入细胞质的示踪剂颗粒。然而，只有一小部分粒子真正进入了原子核。通过进行两次注射实验，还确定了单个毛孔具有双功能，即能够运输蛋白质和RNA。
In the present study, various sized gold particles coated with tRNA, 5S RNA, or poly(A) were used to localize and characterize the pathways for RNA translocation to the cytoplasm. RNA-coated gold particles were microinjected into the nucleus of Xenopus oocytes. The cells were fixed after 15, 60 min, or 6 h, and the particle distribution was later observed by electron microscopy. Similar results were obtained with all classes of RNA used. After nuclear injection, particles ranging from 20-230 .ANG. in diameter were observed within central channels of the nuclear pores and in the cytoplasm immediately adjacent to the pores. Particles of this size would not be expected to diffuse through the pores, suggesting that some form of mediated transport occurred. In addition, it was found that the translocation process is saturable. At least 97% of the pores analyzed appeared to be involved in the translocation process. Gold coated with nonphysiological polynucleotides (poly[I] or poly[dA]) were also translocated. When nuclei were injected with either BSA-, ovalbumin-, polyglutamic acid-, or PVP-coated gold, the particles were essentially excluded from the pores. These results indicate that the accumulation of RNA-gold within the pores and adjacent cytoplasm was not due to non-specific effects. We conclude that the translocation sites for gold particles coated with different classes of RNA are located in the centers of the nuclear pores and that particles at least 230 .ANG. in diameter can cross the envelope. Tracer particles injected into the cytoplasm were observed within the nuclear pores in areas near the site of injection. However, only a small percentage of the particles actually entered the nucleus. It was also determined, by performing double injection experiments, that individual pores are bifunctional, that is, capable of transporting both proteins and RNA.