Deletion of the B-B' and C-C' regions of inverted terminal repeats reduces rAAV productivity but increases transgene expression.
Deletion of the B-B' and C-C' regions of inverted terminal repeats reduces rAAV productivity but increases transgene expression.
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DOI：
10.1038/s41598-017-04054-4
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发表时间：
2017-07-14
影响因子：
4.6
通讯作者：
Wu X
中科院分区：
文献类型：
--
作者：
Zhou Q;Tian W;Liu C;Lian Z;Dong X;Wu X
腺相关病毒（AAV）的反向末端重复序列（ITRs）对病毒基因组的拯救、复制、包装和整合至关重要。虽然ITR突变在之前的报道中已经被发现，但我们设计了一个新的截断ITR，缺少B-B ‘和C-C ’区域，命名为ITRΔBC，并研究了它对重组AAV （rAAV）病毒基因组复制、包装和表达的影响。比较了ITRΔBC rAAV与野生型（wt） ITR rAAV的包装能力。我们的研究结果显示ITRΔBC rAAV的生产力降低了4倍，这与ITRΔBC rAAV的病毒基因组DNA复制比wt ITR rAAV减少了8倍是一致的。令人惊讶的是，ITRΔBC rAAV的转基因表达明显更高。通过抑制和降解ataxia毛细血管扩张突变（ATM）蛋白和Mre11复合物（MRN），对其潜在机制进行了初步探索，因为ATM和/或MRN通过与wt ITRs的顺式相互作用或结合抑制了rAAV的表达。我们证明了对ITRΔBC rAAV表达的抑制作用减弱。本研究提示ITR缺失可影响AAV的转基因表达，为通过ITR修饰提高AAV表达提供了新的途径。
Inverted terminal repeats (ITRs) of the adeno-associated virus (AAV) are essential for rescue, replication, packaging, and integration of the viral genome. While ITR mutations have been identified in previous reports, we designed a new truncated ITR lacking the B-B’ and C-C’ regions named as ITRΔBC and investigated its effects on viral genome replication, packaging, and expression of recombinant AAV (rAAV). The packaging ability was compared between ITRΔBC rAAV and wild-type (wt) ITR rAAV. Our results showed the productivity of ITRΔBC rAAV was reduced 4-fold, which is consistent with the 8-fold decrease in the replication of viral genomic DNA of ITRΔBC rAAV compared with wt ITR rAAV. Surprisingly, transgene expression was significantly higher for ITRΔBC rAAV. A preliminary exploration of the underlying mechanisms was carried out by inhibiting and degrading the ataxia telangiectasia mutated (ATM) protein and the Mre11 complex (MRN), respectively, since the rAAV expression was inhibited by the ATM and/or MRN through cis interaction or binding with wt ITRs. We demonstrated that the inhibitory effects were weakened on ITRΔBC rAAV expression. This study suggests deletion in ITR can affect the transgene expression of AAV, which provides a new way to improve the AAV expression through ITRs modification.
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影响因子：
5.4
作者：
Liu, Y;Shevchenko, A;Berk, AJ
通讯作者：
Berk, AJ
DOI：
10.1073/pnas.56.1.86
发表时间：
1966-01-01
影响因子：
11.1
作者：
ROSE, JA;HOGGAN, MD;SHATKIN, AJ
通讯作者：
SHATKIN, AJ
影响因子：
5.4
作者：
Schwartz, Rachel A.;Palacios, Jose Alejandro;Weitzman, Matthew D.
通讯作者：
Weitzman, Matthew D.
影响因子：
5.4
作者：
MCCARTY, DM;PEREIRA, DJ;MUZYCZKA, N
通讯作者：
MUZYCZKA, N
影响因子：
5.4
作者：
Grimm, Dirk;Lee, Joyce S.;Kay, Mark A.
通讯作者：
Kay, Mark A.