Reuter Thorsten, Weissbrich Benedikt, Schneider-Schaulies Sibylle, Schneider-Schaulies Jürgen
Institut für Virologie und Immunbiologie, Julius Maximilians Universität, Würzburg, Germany.
J Virol. 2006 Jun;80(12):5951-7. doi: 10.1128/JVI.02453-05.
In contrast to studies with genetically modified viruses, RNA interference allows the analysis of virus infections with identical viruses and posttranscriptional ablation of individual gene functions. Using RNase III-generated multiple short interfering RNAs (siRNAs) against the six measles virus genes, we found efficient downregulation of viral gene expression in general with siRNAs against the nucleocapsid (N), phosphoprotein (P), and polymerase (L) mRNAs, the translation products of which form the ribonucleoprotein (RNP) complex. Silencing of the RNP mRNAs was highly efficient in reducing viral messenger and genomic RNAs. siRNAs against the mRNAs for the hemagglutinin (H) and fusion (F) proteins reduced the extent of cell-cell fusion. Interestingly, siRNA-mediated knockdown of the matrix (M) protein not only enhanced cell-cell fusion but also increased the levels of both mRNAs and genomic RNA by a factor of 2 to 2.5 so that the genome-to-mRNA ratio was constant. These findings indicate that M acts as a negative regulator of viral polymerase activity, affecting mRNA transcription and genome replication to the same extent.
与使用基因改造病毒的研究不同，RNA干扰能够利用相同病毒对病毒感染进行分析，并对单个基因功能进行转录后切除。我们使用RNase III生成的针对麻疹病毒六个基因的多个小干扰RNA（siRNA），发现针对核衣壳（N）、磷蛋白（P）和聚合酶（L）mRNA的siRNA通常能有效下调病毒基因表达，这些mRNA的翻译产物形成核糖核蛋白（RNP）复合体。RNP mRNA的沉默在减少病毒信使RNA和基因组RNA方面非常有效。针对血凝素（H）和融合（F）蛋白mRNA的siRNA降低了细胞间融合的程度。有趣的是，siRNA介导的基质（M）蛋白敲低不仅增强了细胞间融合，还使mRNA和基因组RNA的水平提高了2至2.5倍，从而使基因组与mRNA的比例保持恒定。这些发现表明，M作为病毒聚合酶活性的负调节因子，对mRNA转录和基因组复制的影响程度相同。
