In vitro assembly of prenucleolar bodies in Xenopus egg extract.
In vitro assembly of prenucleolar bodies in Xenopus egg extract.
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DOI：
10.1083/jcb.118.6.1297
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发表时间：
1992-09
期刊：
影响因子：
--
通讯作者：
Scheer U
中科院分区：
文献类型：
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作者：
Bell P;Dabauvalle MC;Scheer U
从纯化的 DNA 或染色质中提取的非洲爪蟾卵提取物中组装的细胞核在许多结构和功能特征上与天然对应物相似。然而，体外重建的细胞核中不存在正常间期细胞核最明显的结构元件——核仁。通过使用针对各种核仁成分的抗体的电镜、细胞学银染色和免疫荧光显微镜，我们发现体外组装的细胞核包含许多不同的聚集体，这些聚集体在几个标准上类似于核仁前体（PNB）。这些PNB样结构的形成需要孔复合物介导的蛋白质核转运，但独立于体外细胞核的遗传内容以及转录和翻译事件。我们的数据表明，体外组装的细胞核能够启动核分裂的早期步骤，但产生的 PNB 无法相互融合，可能是由于缺乏功能性核仁组织者。通过适当的修改，该实验系统应该有助于定义和分析促进 PNB 位点特异性组装成连贯核仁体的条件。
Nuclei assembled in Xenopus egg extract from purified DNA or chromatin resemble their natural counterparts in a number of structural and functional features. However, the most obvious structural element of normal interphase nuclei, the nucleolus, is absent from the in vitro reconstituted nuclei. By EM, cytological silver staining, and immunofluorescence microscopy employing antibodies directed against various nucleolar components we show that nuclei assembled in vitro contain numerous distinct aggregates that resemble prenucleolar bodies (PNBs) by several criteria. Formation of these PNB-like structures requires pore complex-mediated nuclear transport of proteins but is independent of the genetic content of the in vitro nuclei as well as transcriptional and translational events. Our data indicate that nuclei assembled in vitro are capable of initiating early steps of nucleologenesis but that the resulting PNBs are unable to fuse with each other, probably due to the absence of a functional nucleolus organizer. With appropriate modifications, this experimental system should be useful to define and analyze conditions promoting the site- specific assembly of PNBs into a coherent nucleolar body.