Requirement of the protein B23 for nucleolar disassembly induced by the FRGY2a family proteins
Requirement of the protein B23 for nucleolar disassembly induced by the FRGY2a family proteins
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DOI：
10.1074/jbc.m512890200
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发表时间：
2006-03-24
影响因子：
4.8
通讯作者：
Kikyo, N
中科院分区：
文献类型：
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作者：
Gonda, K;Wudel, J;Kikyo, N
在非洲爪蛙体细胞核克隆中，供体核仁在卵子胞质中迅速且几乎完全消失。我们之前已经证明，生殖细胞特异性蛋白FRGY2a和FRGY2b是导致这种异常剧烈的核仁解体的原因。核仁的分解没有抑制前rRNA转录，这是众所周知的核仁分离的触发因素，FRGY2a和FRGY2b的核仁分解机制在很大程度上仍不清楚。在这项研究中，我们寻找FRGY2a相互作用的蛋白质，并通过一系列实验研究它们相互作用的功能后果。我们发现在核仁解体过程中，FRGY2a定位于分离的核仁中，并能够解离纯化的核仁，这表明FRGY2a与核仁成分之间存在直接的相互作用。利用His标签下拉方法，我们确定了丰富而多功能的核仁蛋白B23是FRGY2a及其相关的人类蛋白YB1的潜在靶点。免疫共沉淀法证实FRGY2a/YB1与B23之间存在特异性相互作用。最后，用短干扰RNA敲除B23和随后的加回实验证实了B23是YB1分解核仁所必需的。我们认为FRGY2a和YB1通过隔离B23来分解核仁，B23与前核糖体和其他结构上重要的核仁成分有关。
In Xenopus somatic cell nuclear cloning, the nucleoli of donor nuclei rapidly and almost completely disappear in egg cytoplasm. We previously showed that the germ cell-specific proteins FRGY2a and FRGY2b were responsible for this unusually drastic nucleolar disassembly. The nucleolar disassembly occurs without inhibition of pre-rRNA transcription, a well known trigger for nucleolar segregation, and the mechanism for the nucleolar disassembly by FRGY2a and FRGY2b remains largely unknown. In this study, we searched for FRGY2a-interacting proteins and investigated the functional consequences of their interactions through a series of experiments. We showed that during the nucleolar disassembly, FRGY2a localized to the nucleoli of isolated nuclei and was capable of disassembling purified nucleoli, suggesting a direct interaction between FRGY2a and nucleolar components. Using a His tag pull-down approach, we identified the abundant and multifunctional nucleolar protein B23 as a potential target of FRGY2a and its related human protein YB1. A specific interaction between FRGY2a/YB1 and B23 was confirmed by co-immunoprecipitation. Finally, B23 knockdown using short interfering RNA and a subsequent add-back experiment confirmed that B23 was necessary for nucleolar disassembly by YB1. We propose that FRGY2a and YB1 disassemble nucleoli by sequestering B23, which is associated with pre-ribosomes and other structurally important nucleolar components.