Collaco Roy F, Bevington Joyce M, Bhrigu Vipul, Kalman-Maltese Vivian, Trempe James P
Department of Biochemistry and Cancer Biology, University of Toledo, College of Medicine, Toledo, OH 43614-2598, USA.
Virology. 2009 Sep 15;392(1):24-33. doi: 10.1016/j.virol.2009.06.005. Epub 2009 Jul 23.
During adeno-associated virus and adenovirus (AAV/Ad) coinfection, accumulation of viral genomes and proteins can alter cellular stress responses. To determine how AAV/Ad coinfection affects the host we screened over 60 cellular proteins for their responses. AAV/Ad coinfections induce a robust DNA damage response (DDR) that is distinct from that induced by Ad infection alone. Using chemical inhibitors, deficient cell lines and siRNA knockdowns of the DDR kinases, ATM, ATR and DNA-PK, we determined that DNA-PK and ATM kinases are the initial transducers of this response. AAV/Ad coinfection induces ATM- and DNA-PK mediated phosphorylation of RPA2, NBS1, H2AX and the checkpoint kinases CHK1/2. Inhibition of one or more of the DDR kinases reduces the level of phosphorylation of downstream targets but does not dramatically reduce Ad or AAV protein expression. However, AAV DNA levels are moderately affected by kinase inhibition. These experiments provide new insights into the cellular responses to AAV/Ad coinfections.
在腺相关病毒与腺病毒（AAV/Ad）共感染期间，病毒基因组和蛋白质的积累可改变细胞应激反应。为了确定AAV/Ad共感染如何影响宿主，我们筛选了60多种细胞蛋白以观察它们的反应。AAV/Ad共感染会引发强烈的DNA损伤反应（DDR），这与单独由腺病毒感染引发的反应不同。使用化学抑制剂、DDR激酶缺陷细胞系以及DDR激酶ATM、ATR和DNA-PK的小干扰RNA敲低，我们确定DNA-PK和ATM激酶是这种反应的初始转导因子。AAV/Ad共感染诱导ATM和DNA-PK介导的RPA2、NBS1、H2AX以及检查点激酶CHK1/2的磷酸化。抑制一种或多种DDR激酶会降低下游靶点的磷酸化水平，但不会显著降低腺病毒或AAV蛋白的表达。然而，AAV DNA水平会受到激酶抑制的适度影响。这些实验为细胞对AAV/Ad共感染的反应提供了新的见解。
