Martin F, Schaller A, Eglite S, Schümperli D, Müller B
Abteilung für Entwicklungsbiologie, Zoologisches Institut der Universität Bern, Switzerland.
EMBO J. 1997 Feb 17;16(4):769-78. doi: 10.1093/emboj/16.4.769.
The hairpin structure at the 3' end of animal histone mRNAs controls histone RNA 3' processing, nucleocytoplasmic transport, translation and stability of histone mRNA. Functionally overlapping, if not identical, proteins binding to the histone RNA hairpin have been identified in nuclear and polysomal extracts. Our own results indicated that these hairpin binding proteins (HBPs) bind their target RNA as monomers and that the resulting ribonucleoprotein complexes are extremely stable. These features prompted us to select for HBP-encoding human cDNAs by RNA-mediated three-hybrid selection in Saccharomyces cerevesiae. Whole cell extract from one selected clone contained a Gal4 fusion protein that interacted with histone hairpin RNA in a sequence- and structure-specific manner similar to a fraction enriched for bovine HBP, indicating that the cDNA encoded HBP. DNA sequence analysis revealed that the coding sequence did not contain any known RNA binding motifs. The HBP gene is composed of eight exons covering 19.5 kb on the short arm of chromosome 4. Translation of the HBP open reading frame in vitro produced a 43 kDa protein with RNA binding specificity identical to murine or bovine HBP. In addition, recombinant HBP expressed in S. cerevisiae was functional in histone pre-mRNA processing, confirming that we have indeed identified the human HBP gene.
动物组蛋白mRNA 3'末端的发夹结构控制组蛋白RNA的3'加工、核质运输、翻译以及组蛋白mRNA的稳定性。在核提取物和多核糖体提取物中已鉴定出与组蛋白RNA发夹结合的蛋白质，这些蛋白质在功能上即使不完全相同也存在重叠。我们自己的结果表明，这些发夹结合蛋白（HBP）以单体形式结合其靶RNA，并且所形成的核糖核蛋白复合物极其稳定。这些特性促使我们通过在酿酒酵母中进行RNA介导的三杂交筛选来选择编码HBP的人类cDNA。从一个选定克隆中提取的全细胞提取物含有一种Gal4融合蛋白，该蛋白以类似于富含牛HBP的组分的序列和结构特异性方式与组蛋白发夹RNA相互作用，表明该cDNA编码HBP。DNA序列分析显示，编码序列不包含任何已知的RNA结合基序。HBP基因由八个外显子组成，覆盖4号染色体短臂上的19.5 kb。HBP开放阅读框在体外的翻译产生了一种43 kDa的蛋白质，其RNA结合特异性与小鼠或牛HBP相同。此外，在酿酒酵母中表达的重组HBP在组蛋白前体mRNA加工中具有功能，证实我们确实鉴定出了人类HBP基因。
