Collaborator of alternative reading frame protein (CARF) regulates early processing of pre-ribosomal RNA by retaining XRN2 (5'-3' exoribonuclease) in the nucleoplasm.
Collaborator of alternative reading frame protein (CARF) regulates early processing of pre-ribosomal RNA by retaining XRN2 (5'-3' exoribonuclease) in the nucleoplasm.
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DOI：
10.1093/nar/gkv1069
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发表时间：
2015-12-02
影响因子：
14.9
通讯作者：
Takahashi N
中科院分区：
文献类型：
--
作者：
Sato S;Ishikawa H;Yoshikawa H;Izumikawa K;Simpson RJ;Takahashi N
替代阅读框架蛋白（CARF）的合作者直接与ARF、p53和/或人双微小2蛋白（HDM 2）（一种遍在蛋白连接酶）结合，无需辅因子，并通过形成负反馈循环来调节细胞增殖。尽管ARF、p53和HDM2也参与核糖体生物合成的调节，但CARF在该过程中的参与仍然未被探索。在这项研究中，我们证明CARF与5 ′-3 ′核糖核酸外切酶2（XRN2）相关，XRN2在rRNA的成熟和各种废弃的pre-rRNA物质的降解中起着重要作用。我们发现CARF的过表达增加了XRN2在核质中的定位，并伴随着对前rRNA加工的抑制，导致前rRNA转录物的5 ′-0-1、A0 - 1和E-2片段以及45S/47S前rRNA的5 ′延伸片段在核仁中的积累。XRN 2敲除后也观察到了这一点。通过细胞分级分离和免疫细胞化学分析确定，CARF的敲除增加了核仁组分中XRN 2的量。这些观察结果表明，CARF通过控制XRN2在核质和核仁之间的空间分布来调节核糖体生物发生过程中前rRNA加工的早期步骤。
Collaborator of alternative reading frame protein (CARF) associates directly with ARF, p53, and/or human double minute 2 protein (HDM2), a ubiquitin-protein ligase, without cofactors and regulates cell proliferation by forming a negative feedback loop. Although ARF, p53, and HDM2 also participate in the regulation of ribosome biogenesis, the involvement of CARF in this process remains unexplored. In this study, we demonstrate that CARF associates with 5′-3′ exoribonuclease 2 (XRN2), which plays a major role in both the maturation of rRNA and the degradation of a variety of discarded pre-rRNA species. We show that overexpression of CARF increases the localization of XRN2 in the nucleoplasm and a concomitant suppression of pre-rRNA processing that leads to accumulation of the 5′ extended from of 45S/47S pre-rRNA and 5′-01, A0-1 and E-2 fragments of pre-rRNA transcript in the nucleolus. This was also observed upon XRN2 knockdown. Knockdown of CARF increased the amount of XRN2 in the nucleolar fraction as determined by cell fractionation and by immnocytochemical analysis. These observations suggest that CARF regulates early steps of pre-rRNA processing during ribosome biogenesis by controlling spatial distribution of XRN2 between the nucleoplasm and nucleolus.