Etingov Igor, Pintel David J
Department of Molecular Microbiology and Immunology, University of Missouri-Columbia, School of Medicine, Bond Life Sciences Center, Columbia, Missouri, USA.
J Virol. 2024 Dec 17;98(12):e0088924. doi: 10.1128/jvi.00889-24. Epub 2024 Nov 20.
During infection, the autonomous parvovirus minute virus of mice (MVM) induces cellular DNA breaks and localizes to such sites, which presumably affords an environment beneficial for genome replication. MVM replication also benefits from the DNA damage response (DDR) mediated by the ataxia-telangiectasia mutated (ATM) kinase, while the ataxia telangiectasia and Rad-3 related (ATR) arm of the DDR is disabled, which prevents activation of its primary target, checkpoint kinase 1 (Chk1). We find here that Chk1 inactivation strongly correlates with dephosphorylation of one of its targets, RAD51, known to play a pivotal role in homologous recombination repair (HRR), thus leading to substantial inhibition of DNA repair in infected cells. We demonstrate colocalization of replicating MVM DNA with cellular double-strand breaks (DSBs) during infection, and show that an agent that exogenously induces cellular DSBs significantly increases viral DNA replication levels, establishing a role for cellular genome damage in facilitating virus DNA replication. Additionally, overexpression of active Chk1 during MVM infection was found to re-establish the activating phosphorylation of RAD51 Thr 309, significantly suppress infection-induced reduction of HRR efficiency with a concomitant increase in cellular genome DSBs, and reduce viral DNA replication levels. Thus, we conclude that during infection, MVM inhibition of Chk1 activation enhances viral replication, at least in part, by inhibiting cellular HRR.IMPORTANCEThe autonomous parvovirus minute virus of mice (MVM) has a compact DNA genome encoding a minimum number of proteins. During infection, it induces cellular DNA damage and both utilizes and modifies the subsequent cellular DNA damage response (DDR) in various ways to facilitate its replication. One of MVM's activities in this regard is to inhibit one of the primary arms of the DDR, the ataxia telangiectasia and Rad-3 related (ATR) pathway, which prevents activation of checkpoint kinase 1 (Chk1), a key protein involved in controlling the cellular DDR and preserving genome integrity. We show that prevention by MVM of Chk1 activation leads to inhibition of homologous recombination repair (HRR) of cellular DNA, which helps sustain viral replication. This work illuminates another way in which autonomous parvoviruses adjust the cellular environment for their replicative advantage.
在感染过程中，自主型细小病毒小鼠微小病毒（MVM）会诱导细胞DNA断裂并定位于这些位点，这大概为基因组复制提供了一个有利的环境。MVM的复制还受益于由共济失调毛细血管扩张症突变（ATM）激酶介导的DNA损伤反应（DDR），而DDR中共济失调毛细血管扩张症和Rad-3相关（ATR）分支则被禁用，这会阻止其主要靶点检查点激酶1（Chk1）的激活。我们在此发现，Chk1失活与其一个靶点RAD51的去磷酸化密切相关，已知RAD51在同源重组修复（HRR）中起关键作用，从而导致感染细胞中的DNA修复受到显著抑制。我们证明了在感染期间，复制中的MVM DNA与细胞双链断裂（DSB）共定位，并表明一种外源性诱导细胞DSB的试剂会显著提高病毒DNA复制水平，确立了细胞基因组损伤在促进病毒DNA复制中的作用。此外，发现在MVM感染期间过表达活性Chk1会重新建立RAD51苏氨酸309的激活磷酸化，显著抑制感染诱导的HRR效率降低，同时细胞基因组DSB增加，并降低病毒DNA复制水平。因此，我们得出结论，在感染期间，MVM对Chk1激活的抑制至少部分通过抑制细胞HRR来增强病毒复制。
重要性
自主型细小病毒小鼠微小病毒（MVM）具有一个紧凑的DNA基因组，编码的蛋白质数量最少。在感染过程中，它会诱导细胞DNA损伤，并以各种方式利用和改变随后的细胞DNA损伤反应（DDR）以促进其复制。MVM在这方面的活动之一是抑制DDR的主要分支之一，即共济失调毛细血管扩张症和Rad-3相关（ATR）途径，这会阻止检查点激酶1（Chk1）的激活，Chk1是一种参与控制细胞DDR和维持基因组完整性的关键蛋白质。我们表明，MVM对Chk1激活的阻止会导致细胞DNA同源重组修复（HRR）受到抑制，这有助于维持病毒复制。这项工作揭示了自主型细小病毒为获得复制优势而调节细胞环境的另一种方式。
