Nuclear retention element recruits U1 snRNP components to restrain spliced lncRNAs in the nucleus

In contrast to cytoplasmic localization of spliced mRNAs, many spliced lncRNAs are localized in the nucleus. To investigate the mechanism, we used lncRNA MEG3 as a reporter and mapped a potent nuclear retention element (NRE), deletion of this element led to striking export of MEG3 from the nucleus to the cytoplasm. Insertion of the NRE resulted in nuclear retention of spliced lncRNA as well as spliced mRNA. We further purified RNP assembled on the NRE in vitro and identified the proteins by mass spectrometry. Screen using siRNA revealed depletion of U1 snRNP components SNRPA, SNRNP70 or SNRPD2 caused significant cytoplasmic localization of MEG3 reporter transcripts. Co-knockdown these factors in HFF1 cells resulted in an increased cytoplasmic distribution of endogenous lncRNAs. Together, these data support a model that U1 snRNP components restrain spliced lncRNAs in the nucleus via the interaction with nuclear retention element.

与剪接后mRNA的细胞质定位特征相反，多数剪接后的长链非编码RNA（long non-coding RNA, lncRNA）定位于细胞核内。为探究该现象的分子机制，我们以lncRNA MEG3作为报告转录本，鉴定出一段强效的核滞留元件（Nuclear Retention Element, NRE）；删除该元件后，MEG3会显著从细胞核转运至细胞质。将该NRE序列插入后，无论是剪接后的lncRNA还是剪接后的mRNA，均可实现核滞留。我们进一步体外纯化了结合于该NRE的核糖核蛋白复合物（Ribonucleoprotein, RNP），并通过质谱技术鉴定其蛋白组成。通过小干扰RNA（small interfering RNA, siRNA）筛选实验发现，敲低U1小核糖核蛋白（U1 small nuclear ribonucleoprotein, U1 snRNP）的组分SNRPA、SNRNP70或SNRPD2，会使MEG3报告转录本显著出现细胞质定位。在HFF1细胞中共同敲低这些因子，会导致内源性lncRNA的细胞质分布比例显著升高。综上，上述实验结果支持如下模型：U1 snRNP组分通过与核滞留元件结合，将剪接后的lncRNA限制在细胞核内。