Department of Immunology, Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China.
J Infect. 2013 Oct;67(4):329-41. doi: 10.1016/j.jinf.2013.05.003. Epub 2013 May 16.
To investigate the role of miR-146a in dengue virus (DENV) replication.
Expression levels of miR-146a were measured by real-time PCR and Northern blot. Role of miR-146a was tested by overexpression and inhibition assays. Real-time PCR and 50% tissue culture infective dose (TCID50) assays were used to detect RNA levels and extracellular yields of DENV respectively. Interferon (IFN) levels induced by DENV infection were measured by real-time PCR and ELISA respectively. IFN-β neutralization and RNAi were used to test the involvement of IFN-β in the effects of miR-146a. TNFR-associated factor 6 (TRAF6) level was measured by Western-blot.
miR-146a expression was significantly increased in primary human monocytes and THP-1 cells upon DENV infection. Overexpression of miR-146a increased DENV2 replication, while inhibition of miR-146a decreased the viral replication. miR-146a impaired the IFN production and the DENV2 replication suppressed by miR-146a inhibition was partially restored by neutralization of IFN-β or depletion of interferon receptor (IFNAR) 1 or 2. Furthermore, miR-146a targets TRAF6 and overexpression of TRAF6 reversed the effects of miR-146a on IFN-β induction and viral replication.
DENV infection significantly induced the expression of miR-146a, which facilitated viral replication by targeting TRAF6 and dampening IFN-β production.
研究 miR-146a 在登革病毒（DENV）复制中的作用。
通过实时 PCR 和 Northern blot 测定 miR-146a 的表达水平。通过过表达和抑制测定来检验 miR-146a 的作用。分别使用实时 PCR 和 50%组织培养感染剂量（TCID50）测定来检测 DENV 的 RNA 水平和细胞外产量。分别使用实时 PCR 和 ELISA 来测定 DENV 感染诱导的干扰素（IFN）水平。使用 IFN-β 中和和 RNAi 来测试 IFN-β 在 miR-146a 作用中的参与情况。通过 Western blot 测定 TNFR 相关因子 6（TRAF6）水平。
在原代人单核细胞和 THP-1 细胞中，DENV 感染后 miR-146a 的表达明显增加。miR-146a 的过表达增加了 DENV2 的复制，而 miR-146a 的抑制减少了病毒的复制。miR-146a 抑制了 IFN 的产生，并且 miR-146a 抑制导致的 DENV2 复制部分被 IFN-β 的中和或干扰素受体（IFNAR）1 或 2 的耗竭所恢复。此外，miR-146a 靶向 TRAF6，而过表达 TRAF6 逆转了 miR-146a 对 IFN-β 诱导和病毒复制的作用。
DENV 感染显著诱导了 miR-146a 的表达，其通过靶向 TRAF6 和抑制 IFN-β 产生促进了病毒的复制。
