Kimura Tominori, Hashimoto Iwao, Nishizawa Mikio, Ito Seiji, Yamada Hisao
Laboratory of Microbiology and Cell Biology, Department of Pharmacy, College of Pharmaceutical Sciences, Ritsumeikan University, Kusatsu, Shiga 525-8577, Japan.
Med Mol Morphol. 2010 Sep;43(3):145-57. doi: 10.1007/s00795-010-0492-5. Epub 2010 Sep 21.
We recently reported the chromosome region maintenance 1 (CRM1)-dependent nuclear export of intron-less human interferon-α1 (IFN-α1) mRNA, which encodes a main effecter of host innate immunity. We show that the coding region of IFN-α1 mRNA forms novel secondary structures that are responsible for the CRM1-dependent export of the transcript. Deletion-mutagenesis, in vivo export assays, and computer analyses of the folding potentials of export-competent fragments revealed the presence of a domain, termed the conserved secondary structure (CSS), comprising two adjacent putative stable stem-loop structures (nt 208-452). Internal deletion-mutagenesis and constitutive export assays of each stem-loop structure demonstrated that subregions 308-322 and 352-434 act as a core element by conferring the export function on the CSS. Leptomycin B (LMB) inhibition of the CRM1 pathway decreased the export of core element RNA, implying that the principal site of CRM1 action for exporting IFN-α1 mRNA resides within the core element. An RNPS1 (RNA-binding protein S1, serine-rich domain) cDNA was isolated by yeast three-hybrid screening, using bait containing two CSS regions. We showed that RNPS1 might recognize IFN-α1 mRNP that includes CRM1. The data demonstrate that interaction between RNA structures in the coding region and CRM1 affects the nucleocytoplasmic translocation of IFN-α1 mRNA.
我们最近报道了无内含子的人类干扰素-α1（IFN-α1）mRNA的染色体区域维持蛋白1（CRM1）依赖性核输出，该mRNA编码宿主固有免疫的主要效应因子。我们发现IFN-α1 mRNA的编码区形成了新的二级结构，这些结构负责该转录本的CRM1依赖性输出。缺失诱变、体内输出分析以及对有输出能力片段折叠潜力的计算机分析揭示了一个被称为保守二级结构（CSS）的结构域的存在，它由两个相邻的假定稳定茎环结构（核苷酸208 - 452）组成。每个茎环结构的内部缺失诱变和组成型输出分析表明，子区域308 - 322和352 - 434通过赋予CSS输出功能而作为核心元件起作用。雷帕霉素B（LMB）对CRM1途径的抑制降低了核心元件RNA的输出，这意味着CRM1介导IFN-α1 mRNA输出的主要作用位点位于核心元件内。通过酵母三杂交筛选，使用包含两个CSS区域的诱饵，分离出了一个RNPS1（RNA结合蛋白S1，富含丝氨酸结构域）cDNA。我们发现RNPS1可能识别包含CRM1的IFN-α1 mRNP。这些数据表明编码区RNA结构与CRM1之间的相互作用影响了IFN-α1 mRNA的核质转运。
