Department of Molecular Genetics and Microbiology, College of Medicine, University of Florida, United States; Department of Pediatrics, College of Medicine, University of Florida, United States; UF Genetics Institute, University of Florida, United States.
Department of Pediatrics, College of Medicine, University of Florida, United States; UF Genetics Institute, University of Florida, United States; Powell Gene Therapy Center, College of Medicine, University of Florida, United States.
Virus Res. 2019 Dec;274:197771. doi: 10.1016/j.virusres.2019.197771. Epub 2019 Sep 29.
We compared the phenotypes of three mutant AAV2 viruses containing mutations in arginine amino acids (R585, R588 and R484) previously shown to be involved in AAV2 heparan sulfate binding. The transduction efficiencies of wild type and mutant viruses were determined in the eye, the brain and peripheral organs following subretinal, striatal and intravenous injection, respectively, in mice and rats. We found that each of the three mutants (the single mutant R585A; the double mutant R585, 588A; and the triple mutant R585, 588, 484A) had a unique phenotype compared to wt and each other. R585A was completely defective for transducing peripheral organs via intravenous injection, suggesting that R585A may be useful for targeting peripheral organs by substitution of peptide ligands in the capsid surface. In the brain, all three mutants displayed widespread transduction, with the double mutant R585, 588A displaying the greatest spread and the greatest number of transduced neurons. The double mutant was also extremely efficient for retrograde transport, while the triple mutant was almost completely defective for retrograde transport. This suggested that R484 may be directly involved in interaction with the transport machinery. Finally, the double mutant also displayed improved transduction of the eye compared to wild type and the other mutants.
我们比较了三种突变型 AAV2 病毒的表型，这些病毒在先前的研究中显示含有精氨酸氨基酸（R585、R588 和 R484）突变，这些突变与 AAV2 与硫酸乙酰肝素结合有关。在分别通过视网膜下腔、纹状体和静脉内注射后，我们在小鼠和大鼠中确定了野生型和突变型病毒在眼部、脑部和外周器官中的转导效率。我们发现，与野生型相比，这三种突变体（单突变体 R585A；双突变体 R585、588A；和三突变体 R585、588、484A）中的每一种都有独特的表型。R585A 完全不能通过静脉内注射转导外周器官，这表明 R585A 可能通过在衣壳表面取代肽配体，用于靶向外周器官。在脑部，所有三种突变体均显示出广泛的转导，双突变体 R585、588A 显示出最大的扩散和最多的转导神经元。双突变体也非常高效地进行逆行转运，而三突变体几乎完全不能进行逆行转运。这表明 R484 可能直接参与与转运机制的相互作用。最后，与野生型和其他突变体相比，双突变体也显示出对眼部的转导改善。
