Impact of VP1-Specific Protein Sequence Motifs on Adeno-Associated Virus Type 2 Intracellular Trafficking and Nuclear Entry
Impact of VP1-Specific Protein Sequence Motifs on Adeno-Associated Virus Type 2 Intracellular Trafficking and Nuclear Entry
复制标题
DOI：
10.1128/jvi.00282-12
复制
发表时间：
2012-09-01
影响因子：
5.4
通讯作者：
Kleinschmidt, Juergen A.
中科院分区：
文献类型：
--
作者：
Popa-Wagner, Ruth;Porwal, Manvi;Kleinschmidt, Juergen A.
腺相关病毒2型（Adeno-associated virus type 2，AAV 2）作为一种基因传递载体已引起广泛关注。AAV 2介导的基因转移的标志是病毒衣壳的细胞内构象变化，导致感染相关蛋白结构域的暴露。这些蛋白质结构域位于结构蛋白VP 1和VP 2的N-末端部分，包括催化磷脂酶A（2）结构域和三个碱性氨基酸簇。我们已经确定了额外的蛋白质序列基序位于VP 1/2的N端，也被证明是强制性的病毒感染性。这些基序包括已知参与真核细胞中的蛋白质相互作用、内体分选和信号转导的信号。在不同的AAV血清型中，它们是高度保守的，并且各自基序的关键氨基酸的突变导致严重的感染缺陷表型。特别地，与野生型AAV 2相比，YXXQ序列基序的突变显著减少了病毒衣壳在核周围的积累。有趣的是，AAV 2的细胞内运输显示出与PLA（2）活性无关。此外，突变的三个PDZ结合基序，这是连续位于非常尖端的VP 1 N末端，揭示了核转运缺陷的表型，这表明在核摄取的病毒通过一个迄今未知的机制的作用。
Adeno-associated virus type 2 (AAV2) has gained much interest as a gene delivery vector. A hallmark of AAV2-mediated gene transfer is an intracellular conformational change of the virus capsid, leading to the exposure of infection-relevant protein domains. These protein domains, which are located on the N-terminal portion of the structural proteins VP1 and VP2, include a catalytic phospholipase A(2) domain and three clusters of basic amino acids. We have identified additional protein sequence motifs located on the VP1/2 N terminus that also proved to be obligatory for virus infectivity. These motifs include signals that are known to be involved in protein interaction, endosomal sorting and signal transduction in eukaryotic cells. Among different AAV serotypes they are highly conserved and mutation of critical amino acids of the respective motifs led to a severe infection-deficient phenotype. In particular, mutation of a YXXQ-sequence motif significantly reduced accumulation of virus capsids around the nucleus in comparison to wild-type AAV2. Interestingly, intracellular trafficking of AAV2 was shown to be independent of PLA(2) activity. Moreover, mutation of three PDZ-binding motifs, which are located consecutively at the very tip of the VP1 N terminus, revealed a nuclear transport-defective phenotype, suggesting a role in nuclear uptake of the virus through an as-yet-unknown mechanism.