Infectious Diseases and Microbiology, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
PLoS One. 2011;6(6):e21153. doi: 10.1371/journal.pone.0021153. Epub 2011 Jun 17.
The innate immune responses to virus infection are initiated by either Toll-like receptors (TLR3/7/8/9) or cytoplasmic double-stranded RNA (dsRNA)-recognizing RNA helicases RIG-I and MDA5. To avoid causing injury to the host, these signaling pathways must be switched off in time by negative regulators.
METHODOLOGY/PRINCIPAL FINDINGS: Through yeast-two hybrid screening, we found that an ubiquitin-like protein named protein linking integrin-associated protein to cytoskeleton 1(PLIC-1 or Ubiquilin 1) interacted with the Toll/interleukin-1 receptor (TIR) domain of TLR4. Interestingly, PLIC-1 had modest effect on TLR4-mediated signaling, but strongly suppressed the transcriptional activation of IFN-β promoter through the TLR3-Trif-dependent pathway. Concomitantly, reduction of endogenous PLIC-1 by short-hairpin interfering RNA (shRNA) enhanced TLR3 activation both in luciferase reporter assays as well as in new castle disease virus (NDV) infected cells. An interaction between PLIC-1 and Trif was confirmed in co-immunoprecipitation (Co-IP) and GST-pull-down assays. Subsequent confocal microscopic analysis revealed that PLIC-1 and Trif colocalized with the autophagosome marker LC3 in punctate subcellular structures. Finally, overexpression of PLIC-1 decreased Trif protein abundance in a Nocodazole-sensitive manner.
Our results suggest that PLIC-1 is a novel inhibitor of the TLR3-Trif antiviral pathway by reducing the abundance of Trif.
病毒感染引发的固有免疫反应由 Toll 样受体（TLR3/7/8/9）或细胞质双链 RNA（dsRNA）识别 RNA 解旋酶 RIG-I 和 MDA5 启动。为避免对宿主造成伤害，这些信号通路必须及时被负调控因子关闭。
方法/主要发现：通过酵母双杂交筛选，我们发现一种名为蛋白链接整合素相关蛋白到细胞骨架 1（PLIC-1 或泛素样蛋白 1）的泛素样蛋白与 Toll/白细胞介素-1 受体（TIR）结构域的 TLR4 相互作用。有趣的是，PLIC-1 对 TLR4 介导的信号通路仅有轻微影响，但通过 TLR3-Trif 依赖途径强烈抑制 IFN-β 启动子的转录激活。同时，短发夹干扰 RNA（shRNA）降低内源性 PLIC-1 的表达可增强 TLR3 在荧光素酶报告基因检测以及新城疫病毒（NDV）感染细胞中的激活。PLIC-1 和 Trif 之间的相互作用在共免疫沉淀（Co-IP）和 GST 下拉实验中得到证实。随后的共聚焦显微镜分析显示，PLIC-1 和 Trif 与自噬体标记物 LC3 在点状亚细胞结构中共定位。最后，PLIC-1 的过表达以诺考达唑敏感的方式降低了 Trif 蛋白的丰度。
我们的结果表明，PLIC-1 通过降低 Trif 的丰度，成为 TLR3-Trif 抗病毒通路的一种新型抑制剂。
