Eguchi Akiko, Meade Bryan R, Chang Yung-Chi, Fredrickson Craig T, Willert Karl, Puri Nitin, Dowdy Steven F
Howard Hughes Medical Institute, La Jolla, California, USA.
Nat Biotechnol. 2009 Jun;27(6):567-71. doi: 10.1038/nbt.1541. Epub 2009 May 17.
RNA interference (RNAi) induced by short interfering RNA (siRNA) allows for discovery research and large-scale screening; however, owing to their size and anionic charge, siRNAs do not readily enter cells. Current approaches do not deliver siRNAs into a high percentage of primary cells without cytotoxicity. Here we report an efficient siRNA delivery approach that uses a peptide transduction domain-double-stranded RNA-binding domain (PTD-DRBD) fusion protein. DRBDs bind to siRNAs with high avidity, masking the siRNA's negative charge and allowing PTD-mediated cellular uptake. PTD-DRBD-delivered siRNA induced rapid RNAi in a large percentage of various primary and transformed cells, including T cells, human umbilical vein endothelial cells and human embryonic stem cells. We observed no cytotoxicity, minimal off-target transcriptional changes and no induction of innate immune responses. Thus, PTD-DRBD-mediated siRNA delivery allows efficient gene silencing in difficult-to-transfect primary cell types.
由小干扰RNA（siRNA）诱导的RNA干扰（RNAi）可用于发现研究和大规模筛选；然而，由于其大小和阴离子电荷，siRNAs不易进入细胞。目前的方法在不产生细胞毒性的情况下，无法将siRNAs高效递送至大部分原代细胞中。在此，我们报道了一种高效的siRNA递送方法，该方法使用一种肽转导结构域-双链RNA结合结构域（PTD-DRBD）融合蛋白。DRBDs能以高亲和力结合siRNAs，掩盖siRNA的负电荷，并允许PTD介导的细胞摄取。通过PTD-DRBD递送的siRNA能在大部分不同的原代细胞和转化细胞中快速诱导RNAi，包括T细胞、人脐静脉内皮细胞和人胚胎干细胞。我们未观察到细胞毒性、最小化的脱靶转录变化以及先天免疫反应的诱导。因此，PTD-DRBD介导的siRNA递送能够在难以转染的原代细胞类型中实现高效的基因沉默。
