Interferon-dependent R-loop Induction by Zika Virus Contributes to Growth Attenuation [ZIKV_DENV_IFN_DRIPseq]

Zika virus (ZIKV) infection in human neural progenitors triggers DNA damage and activates DNA damage response, leading to cell cycle arrest that can retard brain development. Here we link the ZIKV- induced S phase arrest to replication fork stalling and R-loop induction. DRIP-seq reveals that ZIKV infection induces R-loops at specific loci strongly enriched in the interferon-stimulated genes (ISGs). Bru-seq results further indicate that nascent ISGs transcripts are prone to R-loop induction upon infection. Knockout of interferon receptor eliminated the R-loops on ISGs and partially rescued S phase arrest in infected cells. And overexpression of RNaseH1 reduced ZIKV-mediated DNA damage and cell cycle arrest. We conclude that unscheduled expression of ISGs induced by ZIKV alters R-loop homeostasis and perturbs replication fork progression, leading to fork stalling and eventually DNA damage. IFN- dependent R-loop induction represents a previously unknown, nucleic acid-based mechanism for cell cycle arrest. Overall design: human glioblastoma SNB-19 cells were treated by MR and PR ZIKV virus, DENV virus or interferon and subjuct for DRIP-seq, untreated Mock control were used for comparison

寨卡病毒（Zika virus, ZIKV）感染人类神经前体细胞后，可引发DNA损伤并激活DNA损伤应答通路，进而导致细胞周期阻滞，该过程会延缓大脑发育。本研究明确了ZIKV诱导的S期细胞周期阻滞与复制叉停滞及R环诱导现象之间的关联。DRIP-seq测序结果显示，ZIKV感染会在干扰素刺激基因（interferon-stimulated genes, ISGs）的特定位点大量诱导R环形成。Bru-seq测序结果进一步表明，感染后新生的ISGs转录本易发生R环形成。敲除干扰素受体可消除ISGs位点上的R环，并部分逆转感染细胞中的S期周期阻滞。过表达RNaseH1则可减轻ZIKV介导的DNA损伤与细胞周期阻滞。本研究得出结论：ZIKV诱导的ISGs异常表达会改变R环稳态，干扰复制叉行进，最终引发复制叉停滞与DNA损伤。依赖干扰素（IFN）的R环诱导现象，代表了一种此前未被发现的、基于核酸的细胞周期阻滞机制。实验整体设计：将人类胶质母细胞瘤SNB-19细胞分别用MR株、PR株ZIKV、登革热病毒（DENV）或干扰素处理，随后进行DRIP-seq测序，并以未处理的Mock对照组作为对照