Sorbonne Universités UPMC Univ Paris 06, Inserm, Institut de Myologie, Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, 105 bd de l'Hôpital, Paris, 13, France.
REGENXBIO, 9600 Blackwell Rd, Rockville, MD, 20850, USA.
Sci Rep. 2018 Jan 9;8(1):210. doi: 10.1038/s41598-017-18604-3.
Adeno-associated virus (AAV) transduction efficiency depends on the way in which cellular proteins process viral genomes in the nucleus. In this study, we have investigated the binding of nuclear proteins to the double stranded D (dsD) sequence of the AAV inverted terminal repeat (ITRs) by electromobility shift assay. We present here several lines of evidence that transcription factors belonging to the RFX protein family bind specifically and selectively to AAV2 and AAV1 dsD sequences. Using supershift experiments, we characterize complexes containing RFX1 homodimers and RFX1/RFX3 heterodimers. Following transduction of HEK-293 cells, the AAV genome can be pulled-down by RFX1 and RFX3 antibodies. Moreover, our data suggest that RFX proteins which interact with transcriptional enhancers of several mammalian DNA viruses, can act as regulators of AAV mediated transgene expression.
腺相关病毒（AAV）的转导效率取决于细胞蛋白在核内处理病毒基因组的方式。在这项研究中，我们通过电泳迁移率变动分析研究了核蛋白与 AAV 反向末端重复（ITR）的双链 D（dsD）序列的结合。我们在这里提供了几条证据，表明属于 RFX 蛋白家族的转录因子特异性且选择性地结合 AAV2 和 AAV1 的 dsD 序列。通过超迁移实验，我们对含有 RFX1 同源二聚体和 RFX1/RFX3 异源二聚体的复合物进行了表征。在 HEK-293 细胞转导后，RFX1 和 RFX3 抗体可将 AAV 基因组拉下。此外，我们的数据表明，与几种哺乳动物 DNA 病毒的转录增强子相互作用的 RFX 蛋白可作为 AAV 介导的转基因表达的调节剂。
