Stutika Catrin, Hüser Daniela, Weger Stefan, Rutz Natalja, Heßler Melanie, Heilbronn Regine
Institute of Virology, Campus Benjamin Franklin, Charité Universitätsmedizin Berlin, Germany.
J Gen Virol. 2015 Apr;96(Pt 4):840-850. doi: 10.1099/vir.0.000034. Epub 2014 Dec 22.
Adeno-associated virus (AAV) type 5 represents the genetically most distant AAV serotype and the only one isolated directly from human tissue. Seroepidemiological evidence suggests herpes simplex virus (HSV) as a helper virus for human AAV5 infections, underlining the in vivo relevance of the AAV-herpesvirus relationship. In this study we analysed, for the first time, HSV helper functions for productive AAV5 replication, and compared these to AAV2. Using a combination of HSV strains and plasmids for individual genes, the previously defined HSV helper functions for AAV2 replication were shown to induce AAV5 gene expression, DNA replication and production of infectious progeny. The helper functions comprise the replication genes for ICP8 (UL29), helicase-primase (UL5/8/52), and DNA polymerase (UL30/42). HSV immediate-early genes for ICP0 and ICP4 further enhanced AAV5 replication, mainly by induction of rep gene expression. In the presence of HSV helper functions, AAV5 Rep co-localized with ICP8 in nuclear replication compartments, and HSV alkaline exonuclease (UL12) enhanced AAV5 replication, similarly to AAV2. UL12, in combination with ICP8, was shown to induce DNA strand exchange on partially double-stranded templates to resolve and repair concatemeric HSV replication intermediates. Similarly, concatemeric AAV replication intermediates appeared to be processed to yield AAV unit-length molecules, ready for AAV packaging. Taken together, our findings show that productive AAV5 replication is promoted by the same combination of HSV helper functions as AAV2.
5型腺相关病毒（AAV）是基因上距离最远的AAV血清型，也是唯一直接从人体组织中分离出来的血清型。血清流行病学证据表明单纯疱疹病毒（HSV）是人类AAV5感染的辅助病毒，突出了AAV与疱疹病毒关系在体内的相关性。在本研究中，我们首次分析了HSV对AAV5有效复制的辅助功能，并将其与AAV2进行比较。使用HSV毒株和单个基因的质粒组合，先前确定的HSV对AAV2复制的辅助功能被证明可诱导AAV5基因表达、DNA复制和感染性子代的产生。辅助功能包括ICP8（UL29）、解旋酶-引物酶（UL5/8/52）和DNA聚合酶（UL30/42）的复制基因。HSV的ICP0和ICP4立即早期基因进一步增强了AAV5的复制，主要通过诱导rep基因表达。在存在HSV辅助功能的情况下，AAV5 Rep与ICP8在核复制区室中共定位，并且HSV碱性核酸外切酶（UL12）增强了AAV5的复制，与AAV2类似。已证明UL12与ICP8结合可在部分双链模板上诱导DNA链交换，以解析和修复串联的HSV复制中间体。同样，串联的AAV复制中间体似乎经过加工以产生AAV单位长度分子，为AAV包装做好准备。综上所述，我们的研究结果表明，与AAV2一样，HSV的相同辅助功能组合可促进AAV5的有效复制。
