Mouse nucleolin binds to 4.5S RNAH, a small noncoding RNA
Mouse nucleolin binds to 4.5S RNAH, a small noncoding RNA
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DOI：
10.1016/j.bbrc.2007.10.117
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发表时间：
2008-01-04
影响因子：
3.1
通讯作者：
Harada, Fumio
中科院分区：
文献类型：
--
作者：
Hirose, Yutaka;Harada, Fumio
4.5S RNAH是啮齿动物特异性的小非编码RNA，与B1短散布元件具有广泛的同源性。尽管已知4.5S RNAH与细胞poly（A）-终止RNA和逆转录病毒基因组RNA相关，但其功能仍不清楚。在这项研究中，我们分析了4.5S RNA结合蛋白在小鼠核提取物中使用凝胶迁移率变化和RNA-蛋白质UV交联测定。我们发现至少有9种不同的多肽（p170、p110、p93、p70、p48、p40、p34、p20和p16.5）在体外与4.5S RNAH特异性相互作用。用抗La抗体鉴定p48为小鼠La蛋白。为了鉴定其他4.5S RNA结合蛋白，我们从小鼠cDNA文库中进行了表达克隆，并获得了来自核仁素mRNA的cDNA克隆。我们使用核仁素特异性抗体鉴定p110为核仁素。使用核仁素的各种缺失突变体的UV交联分析表明，四个串联RNA识别基序中的第三个是4.5S RNAH识别的主要决定因素。从小鼠细胞提取物的亚细胞组分的核仁素的免疫沉淀显示，内源性4.5S RNA H的一部分与核仁素，这个复合物位于核质和核仁。(c)2007爱思唯尔公司All rights reserved.
4.5S RNAH is a rodent-specific small noncoding RNA that exhibits extensive homology to the B1 short interspersed element. Although 4.5S RNAH is known to associate with cellular poly(A)-terminated RNAs and retroviral genomic RNAs, its function remains unclear. In this study, we analyzed 4.5S RNAH-binding proteins in mouse nuclear extracts using gel mobility shift and RNA-protein UV cross-linking assays. We found that at least nine distinct polypeptides (p170, p110, p93, p70, p48, p40, p34, p20, and p16.5) specifically interacted with 4.5S RNAH in vitro. Using anti-La antibody, p48 was identified as mouse La protein. To identify the other 4.5S RNAHbinding proteins, we performed expression cloning from a mouse cDNA library and obtained cDNA clones derived from nucleolin mRNA. We identified p110 as nucleolin using nucleolin- specific antibodies. UV cross-linking analysis using various deletion mutants of nucleolin indicated that the third of four tandem RNA recognition motifs is a major determinant for 4.5S RNAH recognition. Immunoprecipitation of nucleolin from the subcellular fractions of mouse cell extracts revealed that a portion of the endogenous 4.5S RNAH was associated with nucleolin and that this complex was located in both the nucleoplasm and nucleolus. (c) 2007 Elsevier Inc. All rights reserved.